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QuickZyme Total Protein Assay Kit

  • Regulatory status:RUO
  • Type:Colorimetric assay
  • Species:Multispecies
Cat. No. Size Price


Discount QZBTOTPROT1 96 wells (1 kit) $537,77
Discount QZBTOTPROT2 2 x 96 wells (1 kit) $837,61
New QZBTOTPROT5 5 x 96 wells (1 kit) $1472,63
PubMed Product Details
Technical Data

Type

Colorimetric assay

Description

Quantitative measurement of total protein in acid hydrolyzates of fresh, frozen, fixed and paraffin embedded tissues in a fast and simple 96-well plate format!

The QuickZyme total collagen and hydroxyproline assays are frequently used to determine the amount of collagen in biological samples such as tissues, tissue extracts, cell extracts and culture media. For the proper interpretation of the data obtained these should be compared to some reference quantity like wet- or dry- weight of the tissue, amount of protein, DNA etc. Some of these quantities are not easy to measure, or require an additional sample because they cannot be measured directly in acid hydrolyzates. It would be helpful if it was possible to determine such a reference in the acid hydrolyzate also used for determination of total collagen or hydroxyproline, such that no longer additional samples or cumbersome methods would be required. The QuickZyme Total Protein Assay offers a convenient solution in such cases.

In this assay the total amount of amino acids present in the same acid hydrolyzate used in the QuickZyme total collagen and hydroxyproline assays is easily determined. This total amount of amino acids in the hydrolyzate is a direct measure of the amount of protein present in the original sample. Combination of the QuickZyme total protein assay with either the total collagen or hydroxyproline assay allows the expression of data as collagen or hydroxyproline per total amount of protein.

The QuickZyme total protein assay is based on the formation of a blue colored product from genipin with free amino acids (Lee et al. Anal. Chim. Acta 480(2003)267; Fujikawa et al. J. Ferment. Technol. 65(1987)149). The assay measures the total amount of amino acids present in the hydrolyzed sample, with exception of proline and hydroxyproline. The amount of amino acids is a good measure for the amount of protein. A hydrolyzed protein standard is supplied with the kit to directly convert data to mg of protein.

The assay is simple and results in a colored product with an absorbance maximum close to 570 nm. The assay is developed to measure protein in acid tissue and/or protein hydrolyzates in such a way that the assay can be performed directly on the hydrolyzate without a drying or neutralization step. When used in addition to the QuickZyme Total Collagen, or QuickZyme Hydroxyproline assay kits, this assay kit allows for easy expression of the collagen or hydroxyproline amounts per total amount of protein.

Applications

Tissue extract, acid hydrolyzates of tissues, Conditioned media, Cells/cell extracts, Frozen or paraffin tissue sections

Sample Requirements

15 μl

Shipping

At ambient temperature. Upon receipt, store the product at the temperature recommended below.

Storage/Expiration

Unopened kit: Store at 4°C in the dark. Do not use kit components past kit expiration date.

Opened kit / reconstituted reagents: The reconstituted reagents should be stored light protected at 4°C (< 1 week), longtime storage at -20°C (stable for at least 2 months). The diluted color reagent solution should be prepared fresh and used the same day.

Calibration Range

0.00 - 3.00 mg/ml

Limit of Detection

0.007 mg/ml

Summary

Features

  • Quantitative measurement of protein in acid hydrolyzates

  • Species independent

  • Colorimetric read out at 570 nm

  • Broad dynamic range: 0.05 to 3.0 mg/ml. Sensitivity: 0.007 mg/ml

  • Uses BSA as standard

  • Ease-of-use: faster and easier than ELISA

  • Storage at room temperature

Research topic

Extracellular matrix

Summary

Soluble and Insoluble Collagen measurement Collagen is the most abundant protein found in animals. During a healthy life span the insoluble, covalent, cross-linked collagen fibers retain their biophysical functions and shapes. The collagen fibers remain isolated from most of the biochemical activities of the cells. Trauma caused by metabolic internal events, external chemical agents or physical injuries however, quickly lead to dramatic activity, resulting in rapid collagen removal followed by a wound healing response (collagen regeneration and associated remodeling). Newly formed soluble collagen production and the residual insoluble collagen fibers can be monitored using the Sircol Soluble Collagen and Sircol Insoluble Collagen Assays. Collagen proteins contain one or more domains with a triple helical structure. The three chains are described as alpha chains and should not be confused with the alpha helixes found in other proteins. The fibrillar collagens (Types I, II, III, V & XI) have most of their alpha chain structure composed of a continuous repeating tri-peptide sequence made up of glycine in every third amino acid residue [(gly-X-Y)n ]. It is to this sequence that the Sircol Dye binds. Proline is frequently an occupant in the ‘Y’ location of the tri-peptide [(gly-X-Y)n]. Many of these residues are converted, post translation, into hydroxyproline residues prior to triple helix formation and the release of the tropocollagen into the ECM. Until now, an investigator seeking to measure covalent cross-linked insoluble collagen required measurements based on free hydroxyproline content – a procedure that requires strong acid (6.0 M HCl), high temperatures (+ 95 °C) and overnight cooking (18 to 24 hrs).

Summary References (1)

References to Collagen

  • Ricard-Blum S. The collagen family. Cold Spring Harb Perspect Biol. 2011 Jan 1;3(1):a004978. doi: 10.1101/cshperspect.a004978. PMID: 21421911; PMCID: PMC3003457. See more on PubMed
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