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D-Glucose HK Assay Kit

Product code: K-GLUHK-220A

Content:

€285.00

220 assays (manual) / 2200 assays (microplate) / 2000 assays (auto-analyser)

Prices exclude VAT

Available for shipping

North American customers click here
Content: (K-GLUHK-110A)
110 assays (manual) / 1100 assays (microplate) / 1000 assays (auto-analyser)
(K-GLUHK-220A)
220 assays (manual) / 2200 assays (microplate) / 2000 assays (auto-analyser)
Shipping Temperature: Ambient
Storage Temperature: Short term stability: 2-8oC,
Long term stability: See individual component labels
Stability: > 2 years under recommended storage conditions
Analyte: D-Glucose
Assay Format: Spectrophotometer, Microplate, Auto-analyser
Detection Method: Absorbance
Wavelength (nm): 340
Signal Response: Increase
Linear Range: 4 to 80 µg of D-glucose per assay
Limit of Detection: 0.66 mg/L
Reaction Time (min): ~ 5 min
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, MEBAK and ASBC Method Malt 6-D

The D-Glucose HK (Regular) test kit is a high purity reagent for the measurement and analysis of D-glucose in plant and food products. Can be used in combination with other Megazyme's products that require glucose determination.

Note for Content: The number of manual tests per kit can be doubled if all volumes are halved.  This can be readily accommodated using the MegaQuantTM  Wave Spectrophotometer (D-MQWAVE).

Browse more of our monosaccharide and oligosaccharide assay kits.

Scheme-K-GLUHK-220A GLUHK Megazyme

Advantages
  • Extended cofactors stability. Dissolved cofactors stable for > 1 year at 4oC.
  • 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
  • Suitable for manual, microplate and auto-analyser formats
Documents
Certificate of Analysis
Safety Data Sheet
FAQs Assay Protocol Data Calculator Product Performance Validation Report
Publications
Megazyme publication

Measurement of available carbohydrates in cereal and cereal products, dairy products, vegetables, fruit and related food products and animal feeds: First Action 2020.07.

McCleary, B. V. & McLoughlin, C. (2021). Journal of AOAC International, qsab019.

Background: The level of available carbohydrates in our diet is directly linked to two major diseases; obesity and Type II diabetes. Despite this, to date there is no method available to allow direct and accurate measurement of available carbohydrates in human and animal foods. Objective: The aim of this research was to develop a method that would allow simple and accurate measurement of available carbohydrates, defined as non-resistant starch, maltodextrins, maltose, isomaltose, sucrose, lactose, glucose, fructose and galactose. Method: Non-resistant (digestible) starch is hydrolysed to glucose and maltose by pancreatic α-amylase and amyloglucosidase at pH 6.0 with shaking or stirring at 37°C for 4 h. Sucrose, lactose, maltose and isomaltose are completely hydrolyzed by specific enzymes to their constituent monosaccharides, which are then measured using pure enzymes in a single reaction cuvette. Results: A method has been developed that allows the accurate measurement of available carbohydrates in all cereal, vegetable, fruit, food, and feed products, including dairy products. Conclusions: A single-laboratory validation was performed on a wide range of food and feed products. The inter-day repeatability (%RSDr) was <3.58% (w/w) across a range of samples containing 44.1 to 88.9% available carbohydrates. The LOD and LOQ obtained were 0.054% (w/w) and 0.179% (w/w), respectively. The method is all inclusive, specific, robust and simple to use. Highlights: A unique method has been developed for the direct measurement of available carbohydrates, entailing separate measurement of glucose, fructose and galactose; information of value in determining the glycemic index of foods.

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Megazyme publication

Measurement of Starch: Critical evaluation of current methodology.

McCleary, B. V., Charmier, L. M. J. & McKie, V. A. (2018). Starch‐Stärke, 71(1-2), 1800146.

Most commonly used methods for the measurement of starch in food, feeds and ingredients employ the combined action of α‐amylase and amyloglucosidase to hydrolyse the starch to glucose, followed by glucose determination with a glucose oxidase/peroxidase reagent. Recently, a number of questions have been raised concerning possible complications in starch analytical methods. In this paper, each of these concerns, including starch hydrolysis, isomerisation of maltose to maltulose, effective hydrolysis of maltodextrins by amyloglucosidase, enzyme purity and hydrolysis of sucrose and β‐glucans have been studied in detailed. Results obtained for a range of starch containing samples using AOAC Methods 996.11 and 2014 .10 are compared and a new simpler format for starch measurement is introduced. With this method that employs a thermostable α-amylase (as distinct from a heat stable α-amylase) which is both stable and active at 100°C and pH 5.0, 10 samples can be analysed within 2 h, as compared to the 6 h required with AOAC Method 2014.10.

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Megazyme publication

Megazyme “advanced” wine test kits general characteristics and validation.

Charnock, S. J., McCleary, B. V., Daverede, C. & Gallant, P. (2006). Reveue des Oenologues, 120, 1-5.

Many of the enzymatic test kits are official methods of prestigious organisations such as the Association of Official Analytical Chemicals (AOAC) and the American Association of Cereal Chemists (AACC) in response to the interest from oenologists. Megazyme decided to use its long history of enzymatic bio-analysis to make a significant contribution to the wine industry, by the development of a range of advanced enzymatic test kits. This task has now been successfully completed through the strategic and comprehensive process of identifying limitations of existing enzymatic bio-analysis test kits where they occurred, and then using advanced techniques, such as molecular biology (photo 1), to rapidly overcome them. Novel test kits have also been developed for analytes of emerging interest to the oenologist, such as yeast available nitrogen (YAN; see pages 2-3 of issue 117 article), or where previously enzymes were simply either not available, or were too expensive to employ, such as for D-mannitol analysis.

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Megazyme publication
Measurement of carbohydrates in grain, feed and food.

McCleary, B. V., Charnock, S. J., Rossiter, P. C., O’Shea, M. F., Power, A. M. & Lloyd, R. M. (2006). Journal of the Science of Food and Agriculture, 86(11), 1648-1661.

Procedures for the measurement of starch, starch damage (gelatinised starch), resistant starch and the amylose/amylopectin content of starch, β-glucan, fructan, glucomannan and galactosyl-sucrose oligosaccharides (raffinose, stachyose and verbascose) in plant material, animal feeds and foods are described. Most of these methods have been successfully subjected to interlaboratory evaluation. All methods are based on the use of enzymes either purified by conventional chromatography or produced using molecular biology techniques. Such methods allow specific, accurate and reliable quantification of a particular component. Problems in calculating the actual weight of galactosyl-sucrose oligosaccharides in test samples are discussed in detail.

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Megazyme publication

Grape and wine analysis: Oenologists to exploit advanced test kits.

Charnock, S. C. & McCleary, B. V. (2005). Revue des Enology, 117, 1-5.

It is without doubt that testing plays a pivotal role throughout the whole of the vinification process. To produce the best possible quality wine and to minimise process problems such as “stuck” fermentation or troublesome infections, it is now recognised that if possible testing should begin prior to harvesting of the grapes and continue through to bottling. Traditional methods of wine analysis are often expensive, time consuming, require either elaborate equipment or specialist expertise and frequently lack accuracy. However, enzymatic bio-analysis enables the accurate measurement of the vast majority of analytes of interest to the wine maker, using just one piece of apparatus, the spectrophotometer (see previous issue No. 116 for a detailed technical review). Grape juice and wine are amenable to enzymatic testing as being liquids they are homogenous, easy to manipulate, and can generally be analysed without any sample preparation.

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Megazyme publication
Measurement of total starch in cereal products by amyloglucosidase-alpha-amylase method: collaborative study.

McCleary, B. V., Gibson, T. S. & Mugford, D. C. (1997). Journal of AOAC International, 80, 571-579.

An American Association of Cereal Chemists/AOAC collaborative study was conducted to evaluate the accuracy and reliability of an enzyme assay kit procedure for measurement of total starch in a range of cereal grains and products. The flour sample is incubated at 95 degrees C with thermostable alpha-amylase to catalyze the hydrolysis of starch to maltodextrins, the pH of the slurry is adjusted, and the slurry is treated with a highly purified amyloglucosidase to quantitatively hydrolyze the dextrins to glucose. Glucose is measured with glucose oxidase-peroxidase reagent. Thirty-two collaborators were sent 16 homogeneous test samples as 8 blind duplicates. These samples included chicken feed pellets, white bread, green peas, high-amylose maize starch, white wheat flour, wheat starch, oat bran, and spaghetti. All samples were analyzed by the standard procedure as detailed above; 4 samples (high-amylose maize starch and wheat starch) were also analyzed by a method that requires the samples to be cooked first in dimethyl sulfoxide (DMSO). Relative standard deviations for repeatability (RSD(r)) ranged from 2.1 to 3.9%, and relative standard deviations for reproducibility (RSD(R)) ranged from 2.9 to 5.7%. The RSD(R) value for high amylose maize starch analyzed by the standard (non-DMSO) procedure was 5.7%; the value was reduced to 2.9% when the DMSO procedure was used, and the determined starch values increased from 86.9 to 97.2%.

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Publication

Postprandial glycemic and lipidemic effects of black rice anthocyanin extract fortification in foods of varying macronutrient compositions and matrices.

Ou, S. J. L., Yang, D., Pranata, H. P., Tai, E. S. & Liu, M. H. (2023). npj Science of Food, 7(1), 59.

Anthocyanin (ACN) fortification of commonly consumed foods is significant as a dietary strategy against the development of metabolic complications by delivering ACNs at high doses. However, its bioactivity and translated metabolic effects in the presence of varying food matrices and macro-constituents is particularly unclear. This end-to-end study investigates the metabolic effects of black rice ACN extract (BRAE) fortification—from in-vitro enzyme inhibitory activities and digestibility, to downstream in vivo impacts on GI, postprandial glycemia and lipidemia. The in vivo effects were investigated in two separate crossover randomised controlled trials (RCT) of 24 healthy participants each-the first RCT determined the postprandial blood glucose, insulin, and ACN bioavailability to a starch-rich single food over 2 h, while the second RCT determined the postprandial blood glucose, insulin, lipid panel, and lipoprotein particles and subfractions to a starch- and fat-rich composite meal over 4 h. In-vitro findings confirmed the inhibitory activities of major black rice ACNs on carbohydrases (p = 0.0004), lipases (p = 0.0002), and starch digestibility (p < 0.0001). in vivo, a 27-point mean GI reduction of wheat bread was observed with BRAE fortification, despite a non-significant attenuation in postprandial glycemia. Conversely, there were no differences in postprandial glycemia when fortified bread was consumed as a composite meal, but acute lipid profiles were altered: (1) improved plasma HDL-c, ([0.0140 mmol/L, 95% CI: (0.00639, 0.0216)], p = 0.0028), Apo-A1 ([0.0296 mmol/L, 95% CI: (0.00757, 0.0515)], p = 0.0203), and Apo-B ([0.00880 mmol/L, 95% CI: (0.00243, 0.0152)], p = 0.0185), (2) modified LDL and HDL subfractions (p < 0.05), and (3) remodelled lipid distributions in HDL and LDL particles. This end-to-end study indicates the potential of ACN fortification in GI reduction and modulating postprandial lipoprotein profiles to starch- and fat-rich composite meals.

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Publication

Circadian Variation in Human Milk Hormones and Macronutrients.

Suwaydi, M. A., Lai, C. T., Rea, A., Gridneva, Z., Perrella, S. L., Wlodek, M. E. & Geddes, D. T. (2023). Nutrients, 15(17), 3729.

There is an inadequate understanding of the daily variations in hormones and macronutrients in human milk (HM), and sample collection protocols vary considerably from study to study. To investigate changes in these milk components across 24 h, 22 lactating women collected small milk samples before and after each breastfeed or expression from each breast. Test weighing was used to determine the volume of HM consumed in each feed. The concentrations of leptin, adiponectin, insulin, fat, and glucose were measured, and the intakes were calculated. A linear mixed model was fitted to assess within-feed and circadian variation in HM feed volume and concentration, and intakes of several components. The average infant intake of HM was 879 g/24 h. Significantly higher pre-feed concentrations were found for adiponectin and glucose and lower post-feed concentrations were found for insulin and fat. Significant circadian rhythms were displayed for leptin, adiponectin, insulin, glucose (both concentration and intake), fat concentration, and milk volume. These findings demonstrate the necessity for setting up standardised and rigorous sampling procedures that consider both within-feed and circadian variations in HM components to gain a more precise understanding of the impacts of these components on infant health, growth and development.

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Publication

Impact of the solubility of phenolic compounds from highland barley (Hordeum vulgare L.) on their antioxidant property and protein binding affinity.

Qin, W., Wang, Y., Mouhamed, F., Hamaker, B. & Zhang, G. (2023). LWT, 186, 115251.

The impact of the solubility of phenolic compounds (PCs) from highland barley (HB) on their antioxidant property and protein binding affinity was investigated. HB-PCs from the phenolic extract (HBE) were analyzed through the UPLC-QTOF-MS, and five fractions were further divided based on their water solubility. Representative compounds of proanthocyanidins, (+)-catechins, and (−)-epicatechin from water fraction (HB–W, 197.67 ± 4.8 μmol GAE/100 g) were significantly correlated with the antioxidant activity of the HBE. In contrast, the PCs enriched in the acetone fraction (HB-A, 60.2 ± 3.1 μmol GAE/100 g) with a structure of 2-phenyl chromogen ketone group (such as rutin, kaempferol, hesperidin, and quercetin) were positively correlated with the protein binding affinity. Further analysis showed that the water-insoluble HB-A fraction significantly lowered the starch digestibility (37.5%) and postprandial glycemia (17.8%) compared to HB-W fractions. Thus, the solubility of PCs is intimately associated with their biological functions as antioxidants or protein-binding ligands, indicating PCs with strong antioxidant properties or high protein binding affinity are structurally distinct from each other, and the physical properties of phenolic compounds might be an important factor to further the understanding of their health functions and mechanisms.

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Publication

Loss of function of Hog1 improves glycerol assimilation in Saccharomyces cerevisiae

Sone, M., Navanopparatsakul, K., Takahashi, S., Furusawa, C. & Hirasawa, T. (2023). World Journal of Microbiology and Biotechnology, 39(10), 255.

We previously isolated a mutant of Saccharomyces cerevisiae strain 85_9 whose glycerol assimilation was improved through adaptive laboratory evolution. To investigate the mechanism for this improved glycerol assimilation, genome resequencing of the 85_9 strain was performed, and the mutations in the open reading frame of HOG1, SIR3, SSB2, and KGD2 genes were found. Among these, a frameshift mutation in the HOG1 open reading frame was responsible for the improved glycerol assimilation ability of the 85_9 strain. Moreover, the HOG1 gene disruption improved glycerol assimilation. As HOG1 encodes a mitogen-activated protein kinase (MAPK), which is responsible for the signal transduction cascade in response to osmotic stress, namely the high osmolarity glycerol (HOG) pathway, we investigated the effect of the disruption of PBS2 gene encoding MAPK kinase for Hog1 MAPK on glycerol assimilation, revealing that PBS2 disruption can increase glycerol assimilation. These results indicate that loss of function of Hog1 improves glycerol assimilation in S. cerevisiae. However, single disruption of the SSK2, SSK22 and STE11 genes encoding protein kinases responsible for Pbs2 phosphorylation in the HOG pathway did not increase glycerol assimilation, while their triple disruption partially improved glycerol assimilation in S. cerevisiae. In addition, the HOG1 frameshift mutation did not improve glycerol assimilation in the STL1-overexpressing RIM15 disruptant strain, which was previously constructed with high glycerol assimilation ability. Furthermore, the effectiveness of the HOG1 disruptant as a bioproduction host was validated, indicating that the HOG1 CYB2 double disruptant can produce L-lactic acid from glycerol.

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Publication

Characterizing and utilizing oxygen-dependent promoters for efficient dynamic metabolic engineering.

Wichmann, J., Behrendt, G., Boecker, S. & Klamt, S. (2023). Metabolic Engineering, 77, 199-207.

Promoters adjust cellular gene expression in response to internal or external signals and are key elements for implementing dynamic metabolic engineering concepts in fermentation processes. One useful signal is the dissolved oxygen content of the culture medium, since production phases often proceed in anaerobic conditions. Although several oxygen-dependent promoters have been described, a comprehensive and comparative study is missing. The goal of this work is to systematically test and characterize 15 promoter candidates that have been previously reported to be induced upon oxygen depletion in Escherichia coli. For this purpose, we developed a microtiter plate-level screening using an algal oxygen-independent flavin-based fluorescent protein and additionally employed flow cytometry analysis for verification. Various expression levels and dynamic ranges could be observed, and six promoters (nar-strong, nar-medium, nar-weak, nirB-m, yfiD-m, and fnrF8) appear particularly suited for dynamic metabolic engineering applications. We demonstrate applicability of these candidates for dynamic induction of enforced ATP wasting, a metabolic engineering approach to increase productivity of microbial strains that requires a narrow level of ATPase expression for optimal function. The selected candidates exhibited sufficient tightness under aerobic conditions while, under complete anaerobiosis, driving expression of the cytosolic F1-subunit of the ATPase from E. coli to levels that resulted in unprecedented specific glucose uptake rates. We finally utilized the nirB-m promoter to demonstrate the optimization of a two-stage lactate production process by dynamically enforcing ATP wasting, which is automatically turned on in the anaerobic (growth-arrested) production phase to boost the volumetric productivity. Our results are valuable for implementing metabolic control and bioprocess design concepts that use oxygen as signal for regulation and induction.

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Publication

Synergistic effect of arabinoxylan and (1, 3)(1, 4)-β-glucan reduces the starch hydrolysis rate in wheat flour.

Ying, R., Zhou, T., Xie, H. & Huang, M. (2023). Food Hydrocolloids, 141, 108668.

The magnitude of nutrient content variability in wheat grains differs by region owing to various environmental factors. Arabinoxylan (AX), the main component of the wheat grain cell wall, plays a key role in determining the quality of dough and final flour products. In the present study, AX was extracted from 10 different genotypes of wheat grains originating from various regions. The molecular weight difference was the largest between Kaifeng-AX (78 × 103 g/mol) and Yangzhou-AX (175 × 103 g/mol). Furthermore, molecular, functional, and nutritional properties-including texture, antioxidant, and in vitro digestion simulation properties-of AX and/or (1,3)(1,4)-β-glucan (BG) flour products were analyzed with respect to storage conditions. The results showed that the higher the molecular weight of AX in bread, the lower the hydrolysis rate of starch. Moreover, the bread samples with AX and BG had the lowest hydrolysis rate; the two polysaccharides of the cell wall exhibited a synergistic effect on starch hydrolysis rate. The sustainable synergistic action of BG and AX improved the wrapping of starch particles and markedly delayed the hydrolysis rate of starch.

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Publication

Impact of Starch-Rich Food Matrices on Black Rice Anthocyanin Accessibility and Carbohydrate Digestibility.

Ou, S. J. L., Fu, A. S. & Liu, M. H. (2023). Foods, 12(4), 880.

Anthocyanins reduce starch digestibility via carbohydrase-inhibitory pathways, but food matrix effects during digestion may also influence its enzymatic function. Understanding anthocyanin-food matrix interactions is significant as the efficiency of carbohydrase inhibition relies on anthocyanin accessibility during digestion. Therefore, we aimed to evaluate the influence of food matrices on black rice anthocyanin accessibility in relation to starch digestibility in common settings of anthocyanin consumption-its co-ingestion with food, and consumption of fortified food. Our findings indicate that black rice anthocyanin extracts (BRAE) had reduced intestinal digestibility of bread to a larger extent for the co-digestion of BRAE with bread (39.3%) (4CO), than BRAE-fortified bread (25.9%) (4FO). Overall anthocyanin accessibility was about 5% greater from the co-digestion with bread than fortified bread across all digestion phases. Differences in anthocyanin accessibility were also noted with changes to gastrointestinal pH and food matrix compositions-with up to 10.1% (oral to gastric) and 73.4% (gastric to intestinal) reductions in accessibility with pH changes, and 3.4% greater accessibility in protein matrices than starch matrices. Our findings demonstrate that the modulation of starch digestibility by anthocyanin is a combined result of its accessibility, food matrix composition, and gastrointestinal conditions.

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Publication

Heterologous Expression of CFL1 Confers Flocculating Ability to Cutaneotrichosporon oleaginosus Lipid-Rich Cells.

Donzella, S. & Compagno, C. (2022). Journal of Fungi, 8(12), 1293.

Lipid extraction from microbial and microalgae biomass requires the separation of oil-rich cells from the production media. This downstream procedure represents a major bottleneck in biodiesel production, increasing the cost of the final product. Flocculation is a rapid and cheap system for removing solid particles from a suspension. This natural characteristic is displayed by some microorganisms due to the presence of lectin-like proteins (called flocculins/adhesins) in the cell wall. In this work, we showed, for the first time, that the heterologous expression of the adhesin Cfl1p endows the oleaginous species Cutaneotrichosporon oleaginosus with the capacity of cell flocculation. We used Helm’s test to demonstrate that the acquisition of this trait allows for reducing the time required for the separation of lipid-rich cells from liquid culture by centrifugation without altering the productivity. This improves the lipid production process remarkably by providing a more efficient downstream.

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Publication

Metabolic reprogramming of OPA1-deficient cells.

Dai, W., Wang, Z., Wang, Q. A., Chan, D. & Jiang, L. (2022). Cell Mol. Life Sci., 79(10), 517.

OPA1, a dynamin-related GTPase mutated in autosomal dominant optic atrophy, is essential for the fusion of the inner mitochondrial membrane. Although OPA1 deficiency leads to impaired mitochondrial morphology, the role of OPA1 in central carbon metabolism remains unclear. Here, we aim to explore the functional role and metabolic mechanism of OPA1 in cell fitness beyond the control of mitochondrial fusion. We applied [U-13C]glucose and [U-13C]glutamine isotope tracing techniques to OPA1-knockout (OPA1-KO) mouse embryonic fibroblasts (MEFs) compared to OPA1 wild-type (OPA1-WT) controls. Furthermore, the resulting tracing data were integrated by metabolic flux analysis to understand the underlying metabolic mechanism through which OPA1 deficiency reprograms cellular metabolism. OPA1-deficient MEFs were depleted of intracellular citrate, which was consistent with the decreased oxygen consumption rate in these cells with mitochondrial fission that is not balanced by mitochondrial fusion. Whereas oxidative glucose metabolism was impaired, OPA1-deficient cells activated glutamine-dependent reductive carboxylation and subsequently relied on this reductive metabolism to produce cytosolic citrate as a predominant acetyl-CoA source for de novo fatty acid synthesis. Prevention of cytosolic glutamine reductive carboxylation by GSK321, an inhibitor of isocitrate dehydrogenase 1 (IDH1), largely repressed lipid synthesis and blocked cell proliferation in OPA1-deficient MEFs. Our data support that, when glucose oxidation failed to support lipogenesis and proliferation in cells with unbalanced mitochondrial fission, OPA1 deficiency stimulated metabolic anaplerosis into glutamine-dependent reductive carboxylation in an IDH1-mediated manner.

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Safety Information
Symbol : GHS05, GHS08
Signal Word : Danger
Hazard Statements : H314, H360
Precautionary Statements : P201, P202, P260, P264, P280
Safety Data Sheet
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