Chromgen Solution

TMB Chromogen Solution (for ELISA)

The assay can be performed directly with serum and in this case will measure total fructosamine in serum proteins. Alternatively, it is possible to separate albumin from other serum proteins by Affi-Gel blue chromatography and then to perform the colorimetric fructosamine assay. The assay is not sensitive to the labile Schiff’s bases (aldimines) or the free glucose concentration. However, several oxidizable compounds in serum interfere with the assay, in particular bilirubin and uric acid .

A second historically important assay to measure fructosamine is the thiobarbituric acid assay [90,91]. The assay is based on the condensation of 5-hydroxymethylfurfural with thiobarbituric acid under formation of a yellow reaction product. The 5-hydroxymethylfurfural is liberated from fructosamine during mild acid hydrolysis. The assay is subject to serious interference by glucose [93], thus requiring glucose removal by ultrafiltration prior to performing the assay. Thiobarbituric acid also reacts with many other carbonyl compounds and is not commonly used to measure fructosamine anymore. However, the assay is still used to measure the formation of malonyldialdehyde and similar aldehydes resulting from lipid peroxidation [94].

 

TMB (3,3′, 5,5;-tetramethylbenzidine) chromogen solution may be used as a substrate for horseradish peroxidase (HRP) in ELISAs.

Using TMB
Use this reagent as supplied, no dilution or further preparation is required. In the presence of peroxidase (HRP, POD), TMB will be converted to a blue color that can be read at OD650 nm.

Specifications

TMB Solution
HRP (Horseradish Peroxidase) Substrate
Wet Ice
Chemical Substrate
TMB
Colorimetric

Chromogenic Substrates

Chromogenic substrates offer a number of advantages over traditional media based on pH indicators. A color change is seen in a colony after transportation and hydrolysis, and this reduces the potential number of false positives due to mutation by more than 10-fold, as fewer genes are involved in generating a signal.

Rapid Detection Using Chromogenic Substrates

Chromogenic compounds added to conventional or newly devised media are used for the isolation of indicator bacteria. These chromogenic substances are modified either by enzymes (which are typical for the respective bacteria) or by specific bacterial metabolites. After modification, the chromogenic substance changes its color or fluorescence, thus enabling easy detection of colonies displaying the metabolic activity.

In this way, these substances are used to avoid the need for isolation of pure cultures and confirmatory tests. The time required for the determination of different indicator bacteria can be reduced to between 14 to 18 hours. When necessary, the surface antigens can be selected and used as the pathogen recognition element for confirmation.

Chromgen Solution
Chromgen Solution

Glycated Serum Albumin and AGE Receptors

Stefan W. Vetter, in Advances in Clinical Chemistry, 2015

3.2.3 Fructosamine Colorimetric Assay

Several chromogenic substrates have been used to detect fructosamine. These assays are based on the reduction of a chromogenic compound by fructosamine or similar ketoamine compounds (i.e., Amadori products) that can undergo oxidation. Substrates that have been used include MTT (3-(4,5-dimethylthiazol2-yl)-2,4-diphenyltetrazolium bromide) [83] and related tetrazolium compounds. A very frequently used compound is nitroblue tetrazolium (NBT; 2,2′di-p-nitrophenyl-5,5′-diphenyl-(3,3′-dimethoxy)-4,4′-bisphenyleneditetrazolium) [84].

Chromogenic substrates

The chromogenic substrates used in this test are various sugars that are linked to either o-nitrophenol or p-nitrophenol by a glycosidic bond. Hydrolysis of the bond frees the nitrophenol, which is detected by its yellow colour. Nitrophenyl derivatives of a large variety of d and l sugars are commercially available, such as p-nitrophenyl-α-d-glucopyranoside (for α-d-glucosidase), p-nitrophenyl-β-d-glucopyranoside (for β-d-glucosidase), o-nitrophenyl-β-d-galactopyranoside (for β-d-galactosidase), p-nitrophenyl-α-l-rhamnopyranoside (for α-l-rhamnosidase) etc. Bear in mind that hydrolysis of a chromogenic substrate does not necessarily mean that the glycosidase for the natural substrate is present. For instance, Staphylococcus aureus can hydrolyse o-nitrophenyl-β-d-galactopyranoside (ONPG) but not lactose (see Kennedy and Scarborough, 1967). Thus results should be reported as ‘ONPG hydrolysed’, not ‘β-galactosidase present’.

(Method of Edberg and Kontnick, 1986) Prepare Sorensen’s phosphate buffer from solutions A and B. Solution A contains 35.6-g Na2HPO4·2H2O per litre of distilled water; solution B contains 27.6-g NaH2PO4·H2O per litre of distilled water. Combine 40.5 ml of solution A with 9.5 ml of solution B to give 50 ml of 0.2-M phosphate buffer and verify that the pH is 7.4. Dilute the stock buffer fourfold to give a 0.05-M working buffer.

Dissolve the nitrophenyl glycoside in the working buffer to give a concentration of 0.5 mg/ml. Dispense 0.25-ml portions into a 12 mm×75 mm tube. Inoculate the tube with several colonies of the organism to be tested and incubate in a water bath for 1 h at 35°C. Look for the development of a yellow colour. No colour change should occur in an uninoculated control tube.

DAB Chromogen Concentrate

D3226-005 50ml
EUR 334

AEC Chromogen Kit

abx097196-1Kit 1 Kit
EUR 272

DAB Chromogen Kit

abx097197-1Kit 1 Kit
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DAB Chromogen Concentrate

ACB030 30 ml
EUR 119

DAB Chromogen Concentrate

ACB060 60 ml
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DAB Chromogen Concentrate

ACB125 125 ml
EUR 245

DAB Chromogen Concentrate

ACB500 500 ml
EUR 671

DAB Chromogen Concentrate

ACB999 1000 ml
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AEC Chromogen Concentrate

ACD015 15 ml
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AEC Chromogen Concentrate

ACD030 30 ml
EUR 148

AEC Chromogen Concentrate

ACD125 125 ml
EUR 245

AEC Chromogen Concentrate

ACD500 500 ml
EUR 671

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