β萘基β-麥芽三糖 β-Naphthyl-β-maltotrioside 貨號:O-NAPC3-100 Megazyme中文站

β萘基β-麥芽三糖

英文名:β-Naphthyl-β-maltotrioside

貨號:O-NAPC3-100

規格:100 mg

CAS: N/A
Molecular Formula: C28H38O16
Molecular Weight: 630.6
Purity: 93%

High purity β-Naphthyl-β-maltotrioside for use in research, biochemical enzyme assays and in vitro diagnostic analysis.

For the specific measurement of cellulase (endo-1,4-β-glucanase) in enzyme preparations and fermentation products.

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4-硝基苯基β-麥芽三糖 4-Nitrophenyl-β-maltotrioside 貨號:O-PNPC3-100 Megazyme中文站

4-硝基苯基β-麥芽三糖

英文名:4-Nitrophenyl-β-maltotrioside

貨號:O-PNPC3-100

規格:100 mg

CAS: 66451-58-9
Molecular Formula: C24H35NO18
Molecular Weight: 625.5
Purity: > 98%

High purity 4-Nitrophenyl-β-maltotrioside for use in research, biochemical enzyme assays and in vitro diagnostic analysis.

A substrate for the measurement of β-amylase.

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木聚糖酶[單胞桿菌] endo-1,4-β-Xylanase (Aeromonas punctata) 貨號:E-XYNAP Megazyme中文站

木聚糖酶[單胞桿菌]

英文名:endo-1,4-β-Xylanase (Aeromonas punctata)

貨號:E-XYNAP

規格:40 Units

High purity recombinant endo-1,4-beta-Xylanase (Aeromonas punctata) for use in research, biochemical enzyme assays and in vitro diagnostic analysis.

EC 3.2.1.8 
CAZy Family: GH10

Recombinant. From Aeromonas punctata. 
In 3.2 M ammonium sulphate.

Specific activity: ~ 8.4 U/mg (40oC, pH 6.5 on wheat arabinoxylan).

Store at 4oC.

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果聚糖[HK法]檢測試劑盒 Fructan HK Assay Kit 貨號:K-FRUCHK Megazyme中文站

果聚糖[HK法]檢測試劑盒

英文名:Fructan HK Assay Kit

貨號:K-FRUCHK

規格:50 assays per kit

分析物意義:許多食品如洋蔥和種子中的常見組分

Megazyme檢測試劑盒優點:方法新穎、反應快、試劑穩定 

The Fructan HK test kit is suitable for the specific measurement and analysis of all fructo-oligosaccharides (reducing and non-reducing) and of fructan polysaccharide.

UV-method for the determination of Fructan in foodstuffs,
beverages and other materials

Principle:
(sucrase + maltase)
(1) Sucrose + maltosaccharides + H2O → D-glucose + D-fructose

(exo-inulinase + endo-inulinase)
(2) Fructan + H2O → D-glucose + D-fructose

(hexokinase)
(3) D-Glucose + D-fructose + ATP → G-6-P + F-6-P + ADP

(glucose-6-phosphate dehydrogenase)
(4) G-6-P + NADP+ → gluconate-6-phosphate + NADPH + H+

(phosphoglucose isomerase)
(5) F-6-P ↔ G-6-P

Kit size: 50 assays
Method: Spectrophotometric at 340 nm
Total assay time: ~ 90 min
Detection limit: 1-100% of sample weight
Application examples:
Flours, plant materials (e.g. onion), food products and other materials
Method recognition:
This method is a modification of AOAC Method 999.03 and AACC
Method 32-32.01

Advantages

  • Very cost effective
  • All reagents stable for > 12 months after preparation
  • Fructan kits are available only from Megazyme
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included

Q1. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q2. Sometimes a negative absorbance change is obtained for the blank samples, is this normal? Should the real value (negative absorbance change) or “0” be used in the calculation of results?

Sometimes the addition of the last assay component can cause a small negative absorbance change in the blank samples due to a dilution effect and in such cases it is recommended that the real absorbance values be used in the calculation of results.

Q3. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample,  in the relevant MegaCalc spreadsheet (if available) to Megazyme ([email protected]). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

Q4. How can I work out how much sample to extract and what dilution of my sample should be used in the kit assay?

Where the amount of analyte in a liquid sample is unknown, it is recommended that a range of sample dilutions are prepared with the aim of obtaining an absorbance change in the assay that is within the linear range.
Where solid samples are analysed, the weight of sample per volume of water used for sample extraction/preparation can be altered to suit, as can the dilution of the extracted sample prior to the addition of the assay, as per liquid samples.

Q5. I have some doubts about the appearance/quality of a kit component what should be done?

If there are any concerns with any kit components, the first thing to do is to test the standard sample (control sample) that is supplied with the kit and ensure that the expected value (within the accepted variation) is obtained before testing any precious samples. This must be done using the procedure provided in the kit booklet without any modifications to the procedure. If there are still doubts about the results using the standard sample in the kit then send example results in the MegaCalc spread sheet to your product supplier (Megazyme or your local Megazyme distributor).

Q6. How much sample should be used for the clarification/extraction of my sample?

The volume/weight of sample and total volume of the extract can be modified to suit the sample. This will ultimately be dictated by the amount of analyte of interest in the sample and may require empirical determination. For low levels of analyte the sample:extract volume ratio can be increased (i.e. increase the sample and/or decrease the total extraction volume).

Alternatively, for samples with low concentrations of analyte, a larger sample volume can be added to the kit assay. When altering the sample volume adjust the distilled water volume added to the assay accordingly so that the total assay volume is not altered.

Q7. Can the test kit be used to measure biological fluids and what sample preparation method should be used?

The kit assay may work for biological fluids assuming that inositol is present above the limit of detection for the kit after any sample preparation (if required). Centrifugation of the samples and use of the supernatant directly in the kit assay (with appropriate dilution in distilled water) may be sufficient. However, if required a more stringent sample preparation method may be required and examples are provided at the following link:http://www.megazyme.com/docs/analytical-applications-downloads/biological_samples_111109.pdf?sfvrsn=2

The test kit has not been tested using biological fluids as samples because it is not marketed or registered as a medical device. This will therefore require your own validation.

Q8. Can the manual assay format be scaled down to a 96-well microplate format?

The majority of the Megazyme test kits are developed to work in cuvettes using the manual assay format, however the assay can be converted for use in a 96-well microplate format. To do this the assay volumes for the manual cuvette format are reduced by 10-fold. The calculation of results for the manual assay format uses a 1 cm path-length, however the path-length in the microplate is not 1 cm and therefore the MegaCalc spreadsheet or the calculation provided in the kit booklet for the manual format cannot be used for the micropalate format unless the microplate reader being used can.

There a 3 main methods for calculation of results using the microplate format:

  1. The easiest method is to use a microplate reader that has a path-length conversion capability (i.e. the microplater reader can detect the path-length of each well and convert the individual readings to a 1 cm path-length). This will allow values to be calculated using the MegaCalc calculation software which can be found where the product is located on the Megazyme website.
  2. Perform a standard curve of the analyte on each microplate that contains test samples and calculate the result of the test samples from the calibration curve (concentration of analyte versus absorbance).
  3. Perform a standard curve of the analyte in both the cuvette format (i.e. with a 1 cm path-length) and the 96-well microplate format and use these results to obtain a mean conversion factor between the cuvette values and the microplate values. Subsequent assays in the microplate format can then be converted from the calculated conversion factor.

Q9. Can the sensitivity of the kit assay be increased?

For samples with low concentrations of analyte the sample volume used in the kit assay can be increased to increase sensitivity. When doing this the water volume is adjusted to retain the same final assay volume. This is critical for the manual assay format because the assay volume and sample volume are used in the calculation of results.

Q10. When using this kit for quantitative analysis what level of accuracy and repeatability can be expected?

The test kit is extremely accurate – at Megazyme the quality control criteria for accuracy and repeatability is to be within 2% of the expected value using pure analytes.

However, the level of accuracy is obviously analyst and sample dependent.

Q11. Must the minimum absorbance change for a sample always be at least 0.1?

No. The 0.1 change of absorbance is only a recommendation. The lowest acceptable change in absorbance can is dictated by the analyst and equipment (i.e. pipettes and spectrophotometer) and therefore can be can be determined by the user. With accurate pipetting, absorbance changes as low as 0.02 can be used accurately.
If a change in absorbance above 0.1 is required but cannot be achieved due to low concentrations of analyte in a sample, this can be overcome by using a larger sample volume in the assay to increase the absorbance change and thereby increase sensitivity of the assay. When doing this the increased volume of the sample should be subtracted from the distilled water volume that is added to the assay so that the total assay volume is unaltered. The increase sample volume should also be accounted for when calculating final results. 

Q12. Can the sensitivity of the kit assay be increased?

Yes. Samples with the lower concentrations of analyte will generate a lower absorbance change. For samples with low concentrations of analyte, a larger sample volume can be used in the assay to increase the absorbance change and thereby increase sensitivity of the assay. When doing this the increased volume of the sample should be subtracted from the distilled water volume that is added to the assay so that the total assay volume is unaltered. The increase sample volume should also be accounted for when calculating final results.

D-葡萄糖[HK法]檢測試劑盒 D-Glucose HK Assay Kit 貨號:K-GLUHK-110A Megazyme中文站

D-葡萄糖[HK法]檢測試劑盒

英文名:D-Glucose HK Assay Kit

貨號:K-GLUHK-110A

規格:110 assays (manual) / 1100 (microplate) / 1000 (auto-analyser)

分析物意義:常見食品組分,在某些情況下非常重要,如糖尿病產品   

Megazyme檢測試劑盒優點:選擇簡單可用的方法,葡萄糖氧化酶/過氧化酶 /己糖激酶/6-磷酸葡萄糖脫氫酶。試劑穩定

High purity reagents for the assay of D-glucose in plant and food products. Can be used in combination with other Megazyme products that require glucose determination. Content:110 assays per kit

UV-method for the determination of D-Glucose in foodstuffs,
beverages and other materials

Principle:
(hexokinase)
(1) D-Glucose + ATP → G-6-P + ADP

(glucose-6-phosphate dehydrogenase)
(2) G-6-P + NADP+ → gluconate-6-phosphate + NADPH + H+

Kit size: (K-GLUHKR)
110 assays (manual) / 1100 (microplate)
/ 1000 (auto-analyser) or
(K-GLUHKL)
220 assays (manual) / 2200 (microplate)
/ 2000 (auto-analyser)
Method: Spectrophotometric at 340 nm
Reaction time: ~ 5 min
Detection limit: 0.66 mg/L
Application examples:
Wine, beer, fruit juices, soft drinks, milk, jam, dietetic foods, bakery
products, candies, fruit and vegetables, tobacco, cosmetics, pharmaceuticals
(e.g. infusions), feed, paper (and cardboard) and other materials (e.g.
biological cultures, samples, etc.)
Method recognition:
Methods based on this principle have been accepted by AOAC, EN,
NEN, NF, DIN, GOST, OIV, IFU, AIJN and MEBAK

Advantages

  • Very competitive price (cost per test)
  • All reagents stable for > 2 years after preparation
  • Rapid reaction
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included
  • Extended cofactors stability
  • Suitable for manual, microplate and auto-analyser formats

 Q1. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q2. Sometimes a negative absorbance change is obtained for the blank samples, is this normal? Should the real value (negative absorbance change) or “0” be used in the calculation of results?

Sometimes the addition of the last assay component can cause a small negative absorbance change in the blank samples due to a dilution effect and in such cases it is recommended that the real absorbance values be used in the calculation of results.

Q3. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample, in the relevant MegaCalc spreadsheet (if available) to Megazyme ([email protected]). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

Q4. Is it possible to detect glucose when it is bound via a glycosidic linkage?

No.  The K-GLUHKL test kit is specific for the measurement of “free” D-glucose.  It will not detect glucose that is bound by a glycosidic linkage to another sugar molecule. 

Q5. Can K-GLUHKL be used to measure glucose in biological samples?

Yes.  It is possible that biological samples may be used directly after appropriate sample dilution in distilled water, however some biological samples may require deproteinisation with perchloric acid prior to addition to the assay.  The deproteinisation method can be found at the following link on the Megazyme website: Deproteinisation Method

Dilution during sample preparation must be taken into account in the final calculation.

Q6. Can K-GLUHKL be used to measure glucose as a component of polysaccharides in plant material?

Yes.  Determination of D-glucose in polysaccharides and fibrous plant material: 
Mill plant material or polysaccharide to pass a 0.5 mm screen using a Retsch centrifugal mill, or similar.  Accurately weigh approx. 100 mg of material into a Corning screw-cap culture tube (16 x 125 mm).  Add 5 mL of 1.3 M HCl to each tube and cap the tubes.  Incubate the tubes at 100?C for 1 h.  Stir the tubes intermittently during the incubation.  Cool the tubes to room temperature, carefully loosen the caps and add 5 mL of 1.3 M NaOH.  Quantitatively transfer the contents of the tube to a 100 mL volumetric flask using distilled water and adjust the volume to 100 mL with distilled water.  Mix thoroughly by inversion and filter an aliquot of the solution through Whatman No. 1 filter paper or centrifuge at 1,500 g for 10 min.  Typically, no further dilution is required and a sample volume of 0.1 mL is satisfactory. 

Q7. Can oligosaccharides or polysaccharides be measured using the kit assay?

The kit assay will only measure the non-covalently linked monosaccharide.

Oligosaccharides or polysaccharides can be measured after hydrolysis to the monosaccharide. Generally acid hydrolysis can be achieved by boiling the oligo/polysaccharide in 1.3 M HCl for 1 h. It is recommended that scientific literature is consulted for information on hydrolysis conditions for the particular oligo/polysaccharide that is being measured.

Q8. Is it possible to add a larger volume then 2 μL of enzyme to the microplate assay? In some instances 2 μL can be difficult to pipette manually.

Yes, instead of adding 2 μL of enzyme suspension an alternative is to dilute the enzyme and add a larger volume to the microplate assay.

Dilute the assay buffer 10-fold with distilled water and use this as the diluent to dilute an aliquot of the enzyme suspension also by 10-fold. Instead of 2 μL, use 20 μL of the diluted enzyme in the microplate assay.

阿拉伯聚糖檢測試劑盒 Arabinan Assay Kit 貨號:K-ARAB Megazyme中文站

阿拉伯聚糖檢測試劑盒

英文名:Arabinan Assay Kit

貨號:K-ARAB

規格:100 assays per kit

The Arabinan test kit is suitable for the measurement and analysis of Arabinan in fruit juice concentrates.

 

 

Advantages

  • Very rapid reaction due to inclusion of galactose mutarotase (patented technology)
  • Very cost effective
  • All reagents stable for > 2 years after preparation
  • Only enzymatic kit available
  • Very specific
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included

 

UV-method for the determination of Arabinan in plant
materials and juices

Principle:
(endo-arabinanase + α-L-arabinofuranosidase)
(1) Arabinan + H2O → L-arabinose

(galactose mutarotase)
(2) α-L-Arabinose ↔ β-L-arabinose

(β-galactose dehydrogenase)
(3) β-L-Arabinose + NAD+ → L-arabinonic acid + NADH + H+

Kit size: 100 assays
Method: Spectrophotometric at 340 nm
Reaction time: ~ 10 min
Detection limit: 1.3 mg/L
Application examples:
Fruit juices and other materials
Method recognition: Novel method

Q1. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q2. I have purchased sugar beet arabinan plus rye and wheat flour arabinoxylans. I would like any information that you could provide on the structures of these polymers.

Sugar Beet Arabinan – this is a polymer of 1,5-α-L-linked arabinofuranose units which is highly substituted by 1,3- and (1-2) linked single a-L-arabinofuranose residues.  About 50% of 1,5 linked arabinosyl residues in the main chain are substituted by 1,3 or 1,2 linked arabinofuranosyl branches.
Rye/Wheat Arabinoxylan – These simply are composed of 1,4-β-D-linked xylan main chains (about 500-1000 residues); substituted by α-L-arabinofuranosyl residues linked 1,3-α or 1,2-α.  They differ in the extent of substitution by α-L-arabinofuranose.  They also contain some ferulic acid residues (linked to arabinose).

Q3. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample,  in the relevant MegaCalc spreadsheet (if available) to Megazyme ([email protected]). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

Q4. What is the molecular weight of Arabinan (Polysaccharide)?

The molecular weight is approximately 15,000 daltons based on HPLC on Fractogel TSK G4000 PN.

天門冬酰苯丙氨酸甲酯/甜味劑/阿斯巴甜[糖精]檢測試劑盒 Aspartame 貨號:K-ASPTM Megazyme中文站

天門冬酰苯丙氨酸甲酯/甜味劑/阿斯巴甜[糖精]檢測試劑盒

英文名:Aspartame

貨號:K-ASPTM

規格:50 assays (manual) /

分析物意義: 食品中常見的增甜劑有阿斯巴甜、D-甘露醇、D-山梨醇和木糖醇

Megazyme檢測試劑盒優點:K-ASPTM-方法新穎/只有試劑盒可用

The Aspartame test kit is a simple and reliable method for the specific measurement and analysis of Aspartame in beverages and foodstuffs.

Suitable for manual, auto-analyser and microplate formats.

UV-method for the determination of Aspartame (and breakdown 
products) in foodstuffs, beverages and other materials

Principle:
                       (pH 12.5)
(1) Asp-Phe-O-Me → Asp-Phe + MeOH

                      (dipeptidase M)
(2) Asp-Phe + H2O → L-aspartate + L-phenylalanine

                      (glutamate-oxaloacetate transaminase)
(3) L-Aspartate + 2-oxoglutarate → L-glutamate + oxaloacetate

                                 (L-malate dehydrogenase)
(4) Oxaloacetate + NADH + H+ → L-malate + NAD+

Kit size:                            50 assays (manual) / 500 (microplate)
                                         / 500 (auto-analyser)
Method:                            Spectrophotometric at 340 nm
Reaction time:                  ~ 5 min
Detection limit:                 0.57 mg/L
Application examples:
Soft drinks, artificial sweeteners, candies, mints, chewing gum, dietetic
products, jam, chocolate and other materials
Method recognition:      Novel method

Advantages

  • Very cost effective
     
  • All reagents stable for > 12 months after preparation
     
  • Only enzymatic kit available
     
  • Measures aspartame and breakdown products (L-aspartate and aspartame acid)
     
  • Very specific
     
  • Very rapid reaction
     
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
     
  • Standard included
     
  • Suitable for manual, microplate and auto-analyser formats

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Alpha淀粉酶檢測底物 Amylazyme – 200 Tablets 貨號:T-AMZ-200T Megazyme中文站

Alpha淀粉酶檢測底物

英文名:Amylazyme – 200 Tablets

貨號:T-AMZ-200T

規格:200

High purity dyed and crosslinked Amylazyme tablets for the measurement of enzyme activity, for research, biochemical enzyme assays and in vitro diagnostic analysis.

AACC Method 22-05.01 and RACI Standard Method. For the assay of cereal and microbial α-amylase. Containing AZCL-Amylose. Recommended substrate for the assay of α-amylase in weather-damaged cereal grains, honey samples and food products containing low levels of this activity.

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D-葡萄糖醛酸和D-半乳糖醛酸檢測試劑盒 D-Glucuronic/D-Galacturonic Acid Assay Kit 貨號:K-URONIC Megazyme中文站

D-葡萄糖醛酸和D-半乳糖醛酸檢測試劑盒

英文名:D-Glucuronic/D-Galacturonic Acid Assay Kit

貨號:K-URONIC

規格:100 assays (manual) /

The D-Glucuronic/D-Galacturonic test kit is a simple, reliable and accurate method for the measurement and analysis of D-hexuronic acids (specifically D-glucuronic acid and D-galacturonic acid) in plant extracts, culture media/supernatants and other materials.
Suitable for manual, auto-analyser and microplate formats.

UV-method for the determination of D-Glucuronic Acid or
D-Galacturonic Acid in hydrolysates of plant material and
polysaccharides and other materials

Principle:
(Uronate dehydrogenase; UDH)
(1) D-Glucuronic acid + NAD+ + H2O → D-glucarate + NADH + H+

(Uronate dehydrogenase; UDH)
(2) D-Galacturonic acid + NAD+ + H2O → D-galactarate + NADH + H+

Kit size: 100 assays (manual) / 1000 (microplate)
/ 1000 (auto-analyser)
Method: Spectrophotometric at 340 nm
Reaction time: ~ 10 min at 25°C or ~ 5 min at 37°C
Detection limit: ~ 17 mg/L
Application examples:
Hydrolysates of plant material and polysaccharides and other
materials
Method recognition: Novel method

Advantages

  • Very cost effective
  • All reagents stable for > 2 years during use
  • Only test kit available
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included
  • Suitable for manual, microplate and auto-analyser formats

 Q1. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q2. Sometimes a negative absorbance change is obtained for the blank samples, is this normal? Should the real value (negative absorbance change) or “0” be used in the calculation of results?

Sometimes the addition of the last assay component can cause a small negative absorbance change in the blank samples due to a dilution effect and in such cases it is recommended that the real absorbance values be used in the calculation of results.

Q3. Can the K-URONIC kit be used to measure 4-O-methylglucuronic acid as well as D-glucuronic acid?

Yes.  The K-URONIC test kit will measure 4-O-methylglucuronic acid as released by alphaglucuronidase from aldouronic acids and wheat arabinoxylan as well as D-glucuronic acid.

Q4. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample, in the relevant MegaCalc spreadsheet (if available) to Megazyme ([email protected]). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

Q5. Can the K-URONIC test be used to measure D-glucurono-g-lactone?

Yes.  K-URONIC can be used to measure D-glucurono-g-lactone.Sample preparation procedure for samples to be tested for D-glucurono-g-lactone:
Adjust the pH of sample solution to approximately 11 with 2 M NaOH (e.g. add 200 μL of 2 M NaOH to 1 mL of sample) and incubate at approx. 25?C for 5-10 min.
Monitor the pH of the solution with pH test-strips, and adjust if necessary.  Use an aliquot of this solution in the kit assay, with appropriate dilution in distilled water if required.
The dilution effect of NaOH addition and any further dilution should also be accounted for in the calculation, e.g. the value obtained from the standard calculation should be multiplied by 1.2 which is the dilution factor as a result of adding 200 μL of 2 M NaOH to 1 mL of sample.
In this assay D-Glucurono-g-lactone is determined together with any “free” D-glucuronic acid, if present, and is calculated as total D-glucuronic acid.
To determine the amount of D-Glucuron-y-lactone present, the sample must also be tested for “free” D-glucuronic acid only (i.e. tested without sample pre-treatment with NaOH) and this value then subtracted from the Total D-glucuronic acid value obtained above.  D-Glucurono-y-lactone = Total D-glucuronic acid – “free” D-glucuronic acid.
This sample treatment using NaOH is used to convert D-glucurono-g-lactone to D-glucuronic acid and should be performed after any other sample pre-treatments that are required for a given sample as provided in the kit booklet, e.g. decolourisation, deproteinisation etc.

Q6. How can I work out how much sample to extract and what dilution of my sample should be used in the kit assay?

Where the amount of analyte in a liquid sample is unknown, it is recommended that a range of sample dilutions are prepared with the aim of obtaining an absorbance change in the assay that is within the linear range.
Where solid samples are analysed, the weight of sample per volume of water used for sample extraction/preparation can be altered to suit, as can the dilution of the extracted sample prior to the addition of the assay, as per liquid samples.

Q7. I have some doubts about the appearance/quality of a kit component what should be done?

If there are any concerns with any kit components, the first thing to do is to test the standard sample (control sample) that is supplied with the kit and ensure that the expected value (within the accepted variation) is obtained before testing any precious samples. This must be done using the procedure provided in the kit booklet without any modifications to the procedure. If there are still doubts about the results using the standard sample in the kit then send example results in the MegaCalc spread sheet to your product supplier (Megazyme or your local Megazyme distributor).

Q8. How much sample should be used for the clarification/extraction of my sample?

The volume/weight of sample and total volume of the extract can be modified to suit the sample. This will ultimately be dictated by the amount of analyte of interest in the sample and may require empirical determination. For low levels of analyte the sample:extract volume ratio can be increased (i.e. increase the sample and/or decrease the total extraction volume).

Alternatively, for samples with low concentrations of analyte, a larger sample volume can be added to the kit assay. When altering the sample volume adjust the distilled water volume added to the assay accordingly so that the total assay volume is not altered.

Q9. Can the test kit be used to measure biological fluids and what sample preparation method should be used?

The kit assay may work for biological fluids assuming that inositol is present above the limit of detection for the kit after any sample preparation (if required). Centrifugation of the samples and use of the supernatant directly in the kit assay (with appropriate dilution in distilled water) may be sufficient. However, if required a more stringent sample preparation method may be required and examples are provided at the following link:http://www.megazyme.com/docs/analytical-applications-downloads/biological_samples_111109.pdf?sfvrsn=2

The test kit has not been tested using biological fluids as samples because it is not marketed or registered as a medical device. This will therefore require your own validation.

Q10. Can the manual assay format be scaled down to a 96-well microplate format?

The majority of the Megazyme test kits are developed to work in cuvettes using the manual assay format, however the assay can be converted for use in a 96-well microplate format. To do this the assay volumes for the manual cuvette format are reduced by 10-fold. The calculation of results for the manual assay format uses a 1 cm path-length, however the path-length in the microplate is not 1 cm and therefore the MegaCalc spreadsheet or the calculation provided in the kit booklet for the manual format cannot be used for the micropalate format unless the microplate reader being used can.

There a 3 main methods for calculation of results using the microplate format:

  1. The easiest method is to use a microplate reader that has a path-length conversion capability (i.e. the microplater reader can detect the path-length of each well and convert the individual readings to a 1 cm path-length). This will allow values to be calculated using the MegaCalc calculation software which can be found where the product is located on the Megazyme website.
  2. Perform a standard curve of the analyte on each microplate that contains test samples and calculate the result of the test samples from the calibration curve (concentration of analyte versus absorbance).
  3. Perform a standard curve of the analyte in both the cuvette format (i.e. with a 1 cm path-length) and the 96-well microplate format and use these results to obtain a mean conversion factor between the cuvette values and the microplate values. Subsequent assays in the microplate format can then be converted from the calculated conversion factor.

Q11. Can the sensitivity of the kit assay be increased?

For samples with low concentrations of analyte the sample volume used in the kit assay can be increased to increase sensitivity. When doing this the water volume is adjusted to retain the same final assay volume. This is critical for the manual assay format because the assay volume and sample volume are used in the calculation of results.

Q12. When using this kit for quantitative analysis what level of accuracy and repeatability can be expected?

The test kit is extremely accurate – at Megazyme the quality control criteria for accuracy and repeatability is to be within 2% of the expected value using pure analytes.

However, the level of accuracy is obviously analyst and sample dependent.

Q13. Is it possible to add a larger volume then 2 μL of enzyme to the microplate assay? In some instances 2 μL can be difficult to pipette manually.

Yes, instead of adding 2 μL of enzyme suspension an alternative is to dilute the enzyme and add a larger volume to the microplate assay.

Dilute the assay buffer 10-fold with distilled water and use this as the diluent to dilute an aliquot of the enzyme suspension also by 10-fold. Instead of 2 μL, use 20 μL of the diluted enzyme in the microplate assay.

Q14. Must the minimum absorbance change for a sample always be at least 0.1?

No. The 0.1 change of absorbance is only a recommendation. The lowest acceptable change in absorbance can is dictated by the analyst and equipment (i.e. pipettes and spectrophotometer) and therefore can be can be determined by the user. With accurate pipetting, absorbance changes as low as 0.02 can be used accurately.
If a change in absorbance above 0.1 is required but cannot be achieved due to low concentrations of analyte in a sample, this can be overcome by using a larger sample volume in the assay to increase the absorbance change and thereby increase sensitivity of the assay. When doing this the increased volume of the sample should be subtracted from the distilled water volume that is added to the assay so that the total assay volume is unaltered. The increase sample volume should also be accounted for when calculating final results. 

Q15. Can the sensitivity of the kit assay be increased?

Yes. Samples with the lower concentrations of analyte will generate a lower absorbance change. For samples with low concentrations of analyte, a larger sample volume can be used in the assay to increase the absorbance change and thereby increase sensitivity of the assay. When doing this the increased volume of the sample should be subtracted from the distilled water volume that is added to the assay so that the total assay volume is unaltered. The increase sample volume should also be accounted for when calculating final results.

1:3,1:4-β-D-葡聚糖的纖維素(C) 1,3:1,4 ?-Glucotetraose (C) 30mg 1,3:1,4 BETA-Glucotetraose (C) 30mg 貨號:O-BGTETC Megazyme中文站

1:3,1:4-β-D-葡聚糖的纖維素(C) 1,3:1,4 ?-Glucotetraose (C) 30mg

英文名:1,3:1,4 BETA-Glucotetraose (C) 30mg

貨號:O-BGTETC

規格:30 mg

Synonyms: Cellobiosyl-(1?3)-β-D-Cellobiose + Glucosyl-(1?3)-β-D-Cellotriose, 
1,3:1,4-β-D-Glucotetraose C + 1,3:1,4-β-D-Glucotetraose A
CAS: 103762-93-2 
         58484-04-1
Molecular Formula: C24H42O21
Molecular Weight: 666.6
Purity: > 95%

High purity 32-β-D-Cellobiosyl-cellobiose + 33-β-D-Glucosyl-cellotriose mixture for use in research, biochemical enzyme assays and in vitro diagnostic analysis.

These tetrasaccharides are produced on hydrolysis of 1:3,1:4-β-D-glucan by cellulase.

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果聚糖檢測試劑盒 Fructan Assay Kit 貨號:K-FRUC Megazyme中文站

果聚糖檢測試劑盒

英文名:Fructan Assay Kit

貨號:K-FRUC

規格:100 assays per kit

產品名稱:果聚糖檢測試劑盒
英文名稱:Fructan Assay Kit
型號規格:100次
品牌:愛爾蘭Megaeyme公司

果聚糖廣泛分布于植物界。單子葉植物,雙子葉植物和綠藻中均存在。

以分子結構和分子重量區別果聚糖。也可以分為三類:菊粉組,果聚糖組和帶支鏈組。菊粉組主要或專門由果糖基果糖鍵組成。果聚糖組主要或專門由果糖基果糖鍵組成。帶支鏈組由大量的(2→1)和(2→6)果糖基果糖鍵共同組成。

目前存在很多測定植物材料和食品中果聚糖的方法。被廣泛接受的是水解為D-果糖(和D-葡萄糖)后再測定的方法。這存在將蔗糖,D-果糖和D-葡萄糖分別去除或測定的問題,Pontis(1966)已經報道了蔗糖,D-葡萄糖和D-果糖的去除方法,利用蔗糖酶水解蔗糖,再通過氫氧化鈉煮沸破壞隨之產生的D-果糖和D-葡萄糖及原有的單糖。據報道,蔗糖酶對菊粉組低聚果糖的作用是緩慢的,可以通過選擇正確的孵育時間使其失效。通過測定目前使用的純化的酵母蔗糖酶,發現很難。

但是,本試劑盒提供的方法操作簡單,使用標準實驗室設備,并且是精確性,可重復性和專一性都很好。采用的酶是高純度酶,專一性水解蔗糖,淀粉和果聚糖。采用的蔗糖酶快速水解蔗糖但對1-蔗果三糖和其他果聚糖的水解活性可以忽略不計(McCleary和Blakeney,1999)(圖2)。在底物濃度為10 mg/mL時,蔗糖與1-蔗果三糖的相對水解比率為3,800:1。

原理:

蔗糖被蔗糖酶水解為D-葡萄糖和D-果糖。同時,如果樣品中存在淀粉和麥芽多糖通過高純β-淀粉酶,普魯蘭酶和麥芽糖酶聯合作用被水解D-葡萄糖。用堿性***物處理生成的還原糖,將其繼續被還原為糖醇。用稀釋的乙酸將溶液調至中性,并去除過量的***物。果聚糖被純化果聚糖酶(菊粉外切酶)水解為D-葡萄糖和D-果糖,通過PAHBAH法測定生成的還原糖。這個方法方便操作,D-葡萄糖和D-果糖的對應顏色一致。含有半乳糖苷蔗糖寡糖的樣品如(棉子糖)在蔗糖/淀粉酶混合物(酶溶液1)處理前推薦使用α-半乳糖苷酶(來源于黑曲霉)孵育。釋放的單糖通過堿性***物處理去除。如果處理中包括這一步驟,需要在計算中考慮體積的變化(如溶液5的最終體積應用1.15mL代替1.1mL).

 

分析物意義:許多食品如洋蔥和種子中的常見組分

Megazyme檢測試劑盒優點:方法新穎、反應快、試劑穩定 

The Fructan test kit is suitable for the specific measurement and analysis of fructan in plant extracts and food products containing starch, sucrose and other sugars.

Colourimetric method for the determination of Fructan in plant
products, foodstuffs and other materials

Principle:
(sucrase)
(1) Sucrose + H2O → D-glucose + D-fructose

(β-amylase + maltase + pullulanase)
(2) Starch + maltosaccharides + H2O → D-glucose

(borohydride)
(3) D-Glucose + D-fructose → D-sorbitol + D-mannitol
(non-reducing)

(exo-inulinase + endo-inulinase)
(4) Fructan + H2O → D-glucose + D-fructose

(100°C, 6 min)
(5) D-Glucose + D-fructose + PAHBAH → PAHBAH colour complex

Kit size: 100 assays
Method: Spectrophotometric at 410 nm
Total assay time: ~ 90 min
Detection limit: 1-100% of sample weight
Application examples:
Flours, plant materials (e.g. onion), food products and other materials
Method recognition:
AOAC (Method 999.03), AACC (Method 32-32.01) and CODEX
(Type III Method)

Advantages

  • Very cost effective
  • All kit reagents stable for > 2 years after preparation
  • Unaffected by high sucrose / reducing sugar concentrations
  • Fructan kits are only available from Megazyme
  • Simple format
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included

Q1. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q2. Does the fructanase enzyme hydrolyse Neosugar?

Yes.

Q3. When measuring fructan in starch containing samples, i.e. maize, is there a possibility of incomplete starch hydrolysis in step B?

From our experience, the level of beta-amylase/pullulanase etc. added is more than adequate to completely hydrolyse any starch likely to be present.  However, if there is a suspicion of incomplete starch hydrolysis, you can check this by running the hydrolysis with the pullulanase/beta-amylase etc. over a range of incubation times and comparing the results.

Q4. Could you tell me whether this method simply gives a gross measure of the fructan content, or whether it is possible to specifically determine the inulin content of the material?

The method measures inulin and cereal fructans.

Q5. What typical absorbance values are obtained for fructan control and fructose control?

Fructan Control Flour (29.6%) 0.496 0.498
Fructose Control 1.133 1.135.

Q6. What is the precision of the method for measurement of fructans?

Our reproducibility is usually +/- 5 to 7% of the fructan value.

Q7. Do you have statistics for the variability of your diagnostic test kits? I would like to get an idea what typical %RSD values would be for your oligofructan test kit.

With all our kits we can obtain approx. 5% c.v. in interlaboratory studies.  In-house these values are closer to 3%.

Q8. Can I detect the levan content of some plant materials by use of your fructan kit?

The fructanase in the kit will completely hydrolyse the fructans in plants.  These are either inulin type or "branched" type (as in oat or wheat stems).  We do not know if the enzymes would act on pure levan.

Q9. What actually is the factor value (F)? Is it always 54.5 or does it change depending on the fructose content?

The value of 54.5 is the actual amount of fructose standard used.  This amount of fructose will give a certain colour in the PAHBAH assay.
Thus; F = factor = [54.5 (micrograms of fructose)] divided by the absorbance value obtained for 54.5 micrograms of fructose in the PAHBAH method.

Q10. Is it possible to buy Fructanase separate from the kit?

Yes.  The best enzyme to use for analytical purposes is exo-Inulinase (E-EXOIAN).

Q11. We need to analyse sucrose in extracts of chicory roots and the sucrase used in your fructan assay procedure may solve the problem. Do you think that it would be possible to use sucrase instead of invertase in the sucrose assay?

Yes, the sugar enzyme can be used to measure sucrose.  The enzyme is an alpha-glucosidase, so it will also hydrolyse maltose if this is present.

Q12. Is it possible to do the weighing and hot extraction the day before the actual assay?

Extract solutions are best made and analysed the same day (not stored overnight). But if they are stored overnight in a refrigerator, heat them to 80?C, and cool before analysing (to ensure all fructan has re-dissolved).

Q13. Can the fructan method be used to measure inulin in starch-containing products, i.e. Cornflakes?

Yes.

Q14. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample,  in the relevant MegaCalc spreadsheet (if available) to Megazyme ([email protected]). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

Q15. Sucrase is specific for the cleavage of sucrose without hydrolysing any of the other potentially present oligo/polysaccharides (mixed linked glucan, soluble starch, fructan of any size & structure). Is this correct? Do you sell the enzyme separately?

Yes , the sucrase does hydrolyse sucrose only and has no action on the other carbohydrates that you mention.  The “sucrase” enzyme is an alpha-glucosidase which we sell as Alpha-Glucosidase (Maltase) E-MALTS.

Q16. Fructan Assay Procedure “Further treatment of extracts” Step 3, Pg 8 "if sample is cooled, it must be re-heated to 80?C to ensure fructans redissolve". If frozen to –20?C, thawed quickly & re-heated to 80?C, will the fructans re-dissolve completely?

Fructans are very soluble and will redissolve very readily.  They may precipitate on freezing (at least the polymeric portion) but they will easily re-dissolve at 80?C.

D-果糖/D-葡萄糖[液體即用型]檢測試劑盒 D-Fructose /D-Glucose (Liquid Ready Reagent) Assay Kit 貨號:K-FRGLQR Megazyme中文站

D-果糖/D-葡萄糖[液體即用型]檢測試劑盒

英文名:D-Fructose /D-Glucose (Liquid Ready Reagent) Assay Kit

貨號:K-FRGLQR

規格:1100 assays per kit in autoanalyser format

The D-Fructose/D-Glucose (Liquid Ready Reagents) test kit is a rapid, reliable and accurate method for the specific measurement and analysis of D-fructose and D-glucose in wine, beverages, foodstuffs and other materials. Supplied as a "ready to use" liquid stable formulation that is suitable for auto-analyser and microplate formats.
Suitable for auto-analyser and microplate formats.

UV-method suitable for auto-analyser and microplate formats for
the determination of D-Fructose and D-Glucose in foodstuffs,
beverages and other materials

Principle:
(hexokinase)
(1) D-Glucose + ATP → G-6-P + ADP

(hexokinase)
(2) D-Fructose + ATP → F-6-P + ADP

(glucose-6-phosphate dehydrogenase)
(3) G-6-P + NADP+ → gluconate-6-phosphate + NADPH + H+

(phosphoglucose isomerase)
(4) F-6-P ↔ G-6-P

Kit size: 1100 assays (microplate) / 1100 (auto-analyser)
Method: Spectrophotometric at 340 nm
Reaction time: ~ 13 min
Detection limit: 133 mg/L (recommended format)
Application examples:
Wine, beer, fruit juices, soft drinks, milk, jam, honey, dietetic foods,
bread, bakery products, candies, desserts, confectionery, ice-cream,
fruit and vegetables, condiments, tobacco, cosmetics, pharmaceuticals,
paper and other materials (e.g. biological cultures, samples, etc.)
Method recognition:
Methods based on this principle have been accepted by AOAC, EN,
NEN, NF, DIN, GOST, OIV, IFU, AIJN, MEBAK and IOCCC

Advantages

  • PVP incorporated to prevent tannin inhibition
  • “Ready to use” liquid stable formulation
  • Very competitive price (cost per test)
  • All reagents stable for > 2 years
  • Very rapid reaction (~ 13 min)
  • Standard included
  • Suitable for microplate and auto-analyser formats

 Q1. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample,  in the relevant MegaCalc spreadsheet (if available) to Megazyme ([email protected]). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

Q2. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q3. Sometimes a negative absorbance change is obtained for the blank samples, is this normal? Should the real value (negative absorbance change) or “0” be used in the calculation of results?

Sometimes the addition of the last assay component can cause a small negative absorbance change in the blank samples due to a dilution effect and in such cases it is recommended that the real absorbance values be used in the calculation of results.

Q4. The pH of my sample is low (pH ~ 3.0), do I need to adjust this before I use the sample in the kit assay?

The final pH of the kit assay after the sample is added should not change from what it should be (as stated in the kit for the assay buffer). If it does change then the sample will require pH adjustment. In most cases the sample volume being used is low relative to the final assay volume and in this case the pH of the kit assay is unlikely to be affected.

Q5. How can I work out how much sample to extract and what dilution of my sample should be used in the kit assay?

Where the amount of analyte in a liquid sample is unknown, it is recommended that a range of sample dilutions are prepared with the aim of obtaining an absorbance change in the assay that is within the linear range.
Where solid samples are analysed, the weight of sample per volume of water used for sample extraction/preparation can be altered to suit, as can the dilution of the extracted sample prior to the addition of the assay, as per liquid samples.

Q6. Can you explain, step by step, how to follow the method and perform the kit assay?

For users who are not familiar with how to use the Megazyme tests kits then it is recommended that they follow this example, e.g. D-Fructose/D-Glucose Assay kit K-FRUGL (http://secure.megazyme.com/D-Fructose-D-Glucose-Assay-Kit):

1. The kit components are listed on pages 2-3 of the kit booklet.
2. Prepare the kit reagents as described on page 3.
3. For separate measurements of glucose and fructose follow procedure A on page 4.
4. Pipette the volumes listed for water, sample, solution 1 and solution 2 into 3 mL, 1 cm pathlength cuvettes. Duplicate sample assays and duplicate blanks are recommended. Mix the contents of each cuvette by inversion (seal the cuvette using parafilm or a plastic cuvette cap – do not use a finger) then after ~3 min record the first absorbance reading of each cuvette at 340 nm (this is reading A1).
5. Then add suspension 3 and mix the contents of each cuvette by inversion. Incubate for 5 minutes then record the absorbance reading of each cuvette at 340 nm (this is reading A2). NB. It is essential that the reaction is compete. To assess this, record the absorbances at ~ 2 minute intervals and until the absorbance plateaus. A stable absorbance indicates that the reaction is complete. If the absorbance continues to increase then continue to record absorbances until it plateaus and only then record absorbance reading A2.
6. Then add suspension 4 and mix the contents of each cuvette by inversion. Incubate for 5 minutes then take absorbance reading of each cuvette at 340 nm (this is reading A3). NB. As above, assess that the reaction has completed by take subsequent readings at ~2 min intervals.
7. For simple, automated results analysis, input the absorbance readings (A1, A2, A3) for samples and blanks into the K-FRUGL MegaCalc.

To ensure that the assay is working, and being performed correctly it is recommend that the test is performed using the standard sample that is provided with the kit and to obtain the expected values before proceeding to test real samples.
It is recommend that new users also watch this video which highlights how to perform the assays.
Many of the other Megazyme test kits follow a similar format.

Q7. I have some doubts about the appearance/quality of a kit component what should be done?

If there are any concerns with any kit components, the first thing to do is to test the standard sample (control sample) that is supplied with the kit and ensure that the expected value (within the accepted variation) is obtained before testing any precious samples. This must be done using the procedure provided in the kit booklet without any modifications to the procedure. If there are still doubts about the results using the standard sample in the kit then send example results in the MegaCalc spread sheet to your product supplier (Megazyme or your local Megazyme distributor).

Q8. Can oligosaccharides or polysaccharides be measured using the kit assay?

The kit assay will only measure the non-covalently linked monosaccharide.

Oligosaccharides or polysaccharides can be measured after hydrolysis to the monosaccharide. Generally acid hydrolysis can be achieved by boiling the oligo/polysaccharide in 1.3 M HCl for 1 h. It is recommended that scientific literature is consulted for information on hydrolysis conditions for the particular oligo/polysaccharide that is being measured.

Q9. Can the sensitivity of the kit assay be increased?

For samples with low concentrations of analyte the sample volume used in the kit assay can be increased to increase sensitivity. When doing this the water volume is adjusted to retain the same final assay volume. This is critical for the manual assay format because the assay volume and sample volume are used in the calculation of results.

Q10. Can the test kit be used to measure biological fluids and what sample preparation method should be used?

The kit assay may work for biological fluids assuming that inositol is present above the limit of detection for the kit after any sample preparation (if required). Centrifugation of the samples and use of the supernatant directly in the kit assay (with appropriate dilution in distilled water) may be sufficient. However, if required a more stringent sample preparation method may be required and examples are provided at the following link:http://www.megazyme.com/docs/analytical-applications-downloads/biological_samples_111109.pdf?sfvrsn=2

The test kit has not been tested using biological fluids as samples because it is not marketed or registered as a medical device. This will therefore require your own validation.

Q11. Can the manual assay format be scaled down to a 96-well microplate format?

The majority of the Megazyme test kits are developed to work in cuvettes using the manual assay format, however the assay can be converted for use in a 96-well microplate format. To do this the assay volumes for the manual cuvette format are reduced by 10-fold. The calculation of results for the manual assay format uses a 1 cm path-length, however the path-length in the microplate is not 1 cm and therefore the MegaCalc spreadsheet or the calculation provided in the kit booklet for the manual format cannot be used for the micropalate format unless the microplate reader being used can.

There a 3 main methods for calculation of results using the microplate format:

  1. The easiest method is to use a microplate reader that has a path-length conversion capability (i.e. the microplater reader can detect the path-length of each well and convert the individual readings to a 1 cm path-length). This will allow values to be calculated using the MegaCalc calculation software which can be found where the product is located on the Megazyme website.
  2. Perform a standard curve of the analyte on each microplate that contains test samples and calculate the result of the test samples from the calibration curve (concentration of analyte versus absorbance).
  3. Perform a standard curve of the analyte in both the cuvette format (i.e. with a 1 cm path-length) and the 96-well microplate format and use these results to obtain a mean conversion factor between the cuvette values and the microplate values. Subsequent assays in the microplate format can then be converted from the calculated conversion factor.

Q12. How much sample should be used for the clarification/extraction of my sample?

The volume/weight of sample and total volume of the extract can be modified to suit the sample. This will ultimately be dictated by the amount of analyte of interest in the sample and may require empirical determination. For low levels of analyte the sample:extract volume ratio can be increased (i.e. increase the sample and/or decrease the total extraction volume).

Alternatively, for samples with low concentrations of analyte, a larger sample volume can be added to the kit assay. When altering the sample volume adjust the distilled water volume added to the assay accordingly so that the total assay volume is not altered.

Q13. Is it possible to add a larger volume then 2 μL of enzyme to the microplate assay? In some instances 2 μL can be difficult to pipette manually.

Yes, instead of adding 2 μL of enzyme suspension an alternative is to dilute the enzyme and add a larger volume to the microplate assay.

Dilute the assay buffer 10-fold with distilled water and use this as the diluent to dilute an aliquot of the enzyme suspension also by 10-fold. Instead of 2 μL, use 20 μL of the diluted enzyme in the microplate assay.

Q14. When using this kit for quantitative analysis what level of accuracy and repeatability can be expected?

The test kit is extremely accurate – at Megazyme the quality control criteria for accuracy and repeatability is to be within 2% of the expected value using pure analytes.

However, the level of accuracy is obviously analyst and sample dependent.

Q15. Must the minimum absorbance change for a sample always be at least 0.1?

No. The 0.1 change of absorbance is only a recommendation. The lowest acceptable change in absorbance can is dictated by the analyst and equipment (i.e. pipettes and spectrophotometer) and therefore can be can be determined by the user. With accurate pipetting, absorbance changes as low as 0.02 can be used accurately.
If a change in absorbance above 0.1 is required but cannot be achieved due to low concentrations of analyte in a sample, this can be overcome by using a larger sample volume in the assay to increase the absorbance change and thereby increase sensitivity of the assay. When doing this the increased volume of the sample should be subtracted from the distilled water volume that is added to the assay so that the total assay volume is unaltered. The increase sample volume should also be accounted for when calculating final results. 

Q16. Can the sensitivity of the kit assay be increased?

Yes. Samples with the lower concentrations of analyte will generate a lower absorbance change. For samples with low concentrations of analyte, a larger sample volume can be used in the assay to increase the absorbance change and thereby increase sensitivity of the assay. When doing this the increased volume of the sample should be subtracted from the distilled water volume that is added to the assay so that the total assay volume is unaltered. The increase sample volume should also be accounted for when calculating final results.

b半乳糖苷酶[黑曲霉] Beta-Galactosidase (A.niger) 8KU 貨號:E-BGLAN Megazyme中文站

b半乳糖苷酶[黑曲霉]

英文名:Beta-Galactosidase (A.niger) 8KU

貨號:E-BGLAN

規格:8000 Units

High purity beta-Galactosidase (A. niger) for use in research, biochemical enzyme assays and in vitro diagnostic analysis.

EC 3.2.1.23 
CAZy Family: GH35

From Aspergillus niger. Highly purified.

Specific activity: > 200 U/mg (40oC, pH 4.5, p-nitrophenyl β-D-galactoside).

Stable at 4oC for > 4 years.

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乳糖/蔗糖/D-葡萄糖檢測試劑盒 Lactose/Sucrose/D-Glucose Assay Kit 貨號:K-LACSU Megazyme中文站

乳糖/蔗糖/D-葡萄糖檢測試劑盒

英文名:Lactose/Sucrose/D-Glucose Assay Kit

貨號:K-LACSU

規格:100 assays of each per kit

分析物意義:常見加工食品組分,在某些情況下,精確的數值很重要,如 “無乳糖”產品 

Megazyme檢測試劑盒優點:K-LACGAR試劑盒反應快(室溫,5min)、試劑穩定

The Lactose/Sucrose/D-Glucose assay kit is suitable for the measurement and analysis of sucrose, lactose and D-glucose in flour mixtures and other materials.

Colourimetric method for the determination of Lactose, Sucrose
and D-Glucose in foodstuffs, beverages and other materials

Principle:
(invertase)
(1) Sucrose + H2O → D-glucose + D-fructose

(β-galactosidase)
(2) Lactose + H2O → D-glucose + D-galactose

(glucose oxidase)
(3) D-Glucose + H2O + O2 → D-gluconate + H2O2

(peroxidase)
(4) 2H2O2 + p-hydroxybenzoic acid + 4-aminoantipyrine →
quinoneimine + 4H2O

Kit size: 100 assays of each
Method: Spectrophotometric at 510 nm
Reaction time: ~ 60 min
Detection limit: 100 mg/L
Application examples:
Flours, beverages, dairy products, milk, foodstuffs containing milk,
cosmetics, pharmaceuticals and other materials (e.g. biological cultures,
samples, etc.)
Method recognition:
Used and accepted in food analysis

Advantages

  • Very competitive price (cost per test)
  • All reagents stable for > 12 months after preparation
  • Simple format
  • Very specific
  • Rapid reaction
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
  • Standard included

 Q1. Would your kit K-LACSU (Lactose/Sucrose/D-Glucose) allow us to analyse for lactose in dairy by-products?

The K-LACSU kit will allow measurement of lactose in any materials.

Q2. Should the pH of the sample be adjusted even for samples in acidic media?

The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
Samples above 0.1 mL are more likely to affect the pH of the assay solution and   therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

Q3. Can your Lactose/Sucrose/D-Glucose Kit (K-LACSU) be used for measuring sucrose in wort and during fermentation?

The Lactose/Sucrose/D-Glucose Kit should work fine for wort.

Q4. With regard to your Lactose/Sucrose/D-Glucose Kit (K-LACSU), is it possible to measure lactose alone in a mixture of material in which sucrose and glucose could be present?

Yes, it is possible to specifically measure lactose in a mixture of sucrose, lactose and glucose.  Of course, it is also necessary to measure free glucose (i.e. with no added beta-galactosidase).

Q5. Would your kits – K-LACSU (Lactose/Sucrose/D-Glucose) and K-TSTA (Total Starch) allow us to analyse mixed feedstuff for total sugars?

The Lactose/Sucrose/D-Glucose Kit will allow measurement of sucrose, lactose and  glucose (not total sugars).  We can supply reagents separately for sucrose or lactose determination (or Glucose).

Q6. There is an issue with the performance of the kit; the results are not as expected.

If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

  1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
  2. Send the results of the kit standard, blank samples and the results obtained for your sample, in the relevant MegaCalc spreadsheet (if available) to Megazyme ([email protected]). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
  3. State the kit lot number being used (this is found on the outside of the kit box).
  4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
  5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
  6. State the sample type and describe the sample preparation steps if applicable.

Q7. It states on the box that the kit should be stored at 0-5?C. The kit was not shipped at this temperature and the average temperature in our country is 30?C. Can we now use this kit or not?

The kits we supply are stable at room temperature for several months.  Thus we ship at room temperature.  When you get the kit, it should be stored as recommended and it will be stable for several years.  The kit will be perfectly fine at 30?C for the shipping time.

4-甲基 – α-mannotriose 4-Methylumbelliferyl-α-mannotrioside 貨號:O-4MUAM3 Megazyme中文站

4-甲基 – α-mannotriose

英文名:4-Methylumbelliferyl-α-mannotrioside

貨號:O-4MUAM3

規格:20 mg

CAS: 66068-41-5
Molecular Formula: C28H38O18

Molecular Weight: 662.6
Purity: 98%

High purity 4-Methylumbelliferyl-α-mannotrioside for use in research, biochemical enzyme assays and in vitro diagnostic analysis. This is a potential colourimetric substrate for research into β-1,4-mannan degrading enzymes.

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2-氯-4-硝基苯基α-甘露二糖 2-Chloro-4-nitrophenyl-α-mannobioside 貨號:O-CPNPAM2 Megazyme中文站

2-氯-4-硝基苯基α-甘露二糖

英文名:2-Chloro-4-nitrophenyl-α-mannobioside

貨號:O-CPNPAM2

規格:20 mg

CAS: N/A
Molecular Formula: C18H24ClNO13
Molecular Weight: 497.8
Purity: > 95%

High purity 2-Chloro-4-nitrophenyl-α-mannobioside for use in research, biochemical enzyme assays and in vitro diagnostic analysis. This is a potential colourimetric substrate for research into β-1,4-mannan degrading enzymes.

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阿拉伯樹膠四糖[純度>95%][漿] Arabinotetraose (syrup) – 30mg 貨號:O-ATE Megazyme中文站

阿拉伯樹膠四糖[純度>95%][漿]

英文名:Arabinotetraose (syrup) – 30mg

貨號:O-ATE

規格:30 mg

CAS: 190852-24-5
Molecular Formula: C20H34O17
Molecular Weight: 546.5
Purity: > 95%



High purity Arabinotetraose (syrup) for use in research, biochemical enzyme assays and in vitro diagnostic analysis.

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阿魏酸酯酶[丁酸桿菌] Feruloyl esterase (Clostridium thermocellum) 貨號:E-FAEZCT Megazyme中文站

阿魏酸酯酶[丁酸桿菌]

英文名:Feruloyl esterase (Clostridium thermocellum)

貨號:E-FAEZCT

規格:10 Units (~ 1800 U on FAXX)

High purity recombinant Feruloyl esterase (Clostridium thermocellum) for use in research, biochemical enzyme assays and in vitro diagnostic analysis.

EC 3.1.1.73 
CAZy Family: CE1

Recombinant. Feruloyl esterase domain of xylanase XynZ (Xyn10A) from Clostridium thermocellum. 
In 3.2 M ammonium sulphate.

Specific activity: ~ 0.6 U/mg on ethyl ferulate at 50oC and pH 6.0. This enzyme is ~ 180 fold more active on FAXX.

Stability: > 2 years at 4oC.

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半乳糖脫氫酶[土壤原核生物] Galactose dehydrogenase (soil prokaryote) 貨號:E-GALDH Megazyme中文站

半乳糖脫氫酶[土壤原核生物]

英文名:Galactose dehydrogenase (soil prokaryote)

貨號:E-GALDH

規格:200 Units

High purity recombinant Galactose dehydrogenase (soil prokaryote) for use in research, biochemical enzyme assays andin vitro diagnostic analysis.

EC 1.1.1.48

Recombinant from soil procaryote. This recombinant enzyme has been expressed in E. coli and purified by affinity chromatography. Electrophoretically homogeneous (MW 36,659). In 3.2 M ammonium sulphate.

Specific activity: 390 U/mg (25oC, pH 8.6, on galactose).

Stable at 4oC for > 2 years.

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木葡聚糖酶(GH74)(類芽孢sp。) Xyloglucanase (GH74) (Paenibacillus sp.) 貨號:E-XGP74 Megazyme中文站

木葡聚糖酶(GH74)(類芽孢sp。)

英文名:Xyloglucanase (GH74) (Paenibacillus sp.)

貨號:E-XGP74

規格:200 Units at 70°C

High purity recombinant exo-alpha-Sialidase (S. Typhimurium) for use in research, biochemical enzyme assays and in vitro diagnostic analysis. 

EC 3.2.1.18
CAZy Family: GH33

Recombinant. From Salmonella typhimurium. In solution (Tris.HCl / NaCl / EDTA). Hydrolysis of unbranched, non-reducing terminal α-2,3-linked >> α-2,6-linked >> α-2,8-linked N-acetylneuraminic acid (NANA; Neu5Ac) residues from glycoproteins and oligosaccharides of glycoconjugates.
Supplied at ~ 2,500 U/mL. 

Specific activity: ~ 750 U/mg (37oC, pH 7.0 on pNP-α-D-N-acetylneuraminic acid)

Store at 4oC.  

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63-α-D-麥芽三糖,麥芽四糖 63-a-D-maltotriosyl-maltotriitol 貨號:O-MTMTRD Megazyme中文站

63-α-D-麥芽三糖,麥芽四糖

英文名:63-a-D-maltotriosyl-maltotriitol

貨號:O-MTMTRD

規格:100mg

Synonym: Borohydride reduced 63-α-D-maltotriosyl-maltotriose
CAS: 107123-59-1
Molecular Formula: C36H64O31
Molecular Weight: 992.9
Purity: > 95%

High purity 63-α-D-maltotriosyl-maltotriitol for use in research, biochemical enzyme assays and in vitro diagnostic analysis. This compound can be used as a substrate for the characeterisation of pullulanase or limit dextrinase enzymes using a reducing sugar method (e.g. Nelson-Somogyi). The borohydride reduced analogue is used in place of 63-α-D-maltotriosyl-maltotriose (Cat. No. O-MTMT), because being a reducing sugar itself, the native oligosaccharide generates a large ‘blank’ value in this assay.

 

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