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quantify collagen in various sources such as cell media, ECM (Extra Cellular Matrix) of culture cell and tissue |
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Human collagen type1 ELISA KIT |
Back Ground
 Type
1 collagen is the most abundant protein in the connective tissues,
especially in tendon, skin and bone. This kit is designed to
quantify collagen in various sources such as cell media, ECM
(Extra Cellular Matrix) of culture cell and tissue because the
kit detects atelo-collagen which is prepared by pepsin digestion.
Features
| Short assay time (2 hours 15 minutes). |
| Collagen pre-coated microtiter-plate. |
| Simultaneous assay of many samples (Assay maximum is 40 samples per 1 kit in duplicate). |
| No need of special machines and equipments because of non-isotope assay. |
| Partitional use because of split type (8 wells/strip). |
Principle(Competitive EIA)
This assay is a competitive EIA in which polyclonal antibody to human type 1 atelo-collagen is used.
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Assay Process: (Assay time 2 hours 15 minutes)
Mixture of sample contained atelo-collagen and biotinylated anti-atelo-collagen antibody is added to a well of microtiter-plate on which purified atelo-collagen is immobilized.
After washing, add peroxidase labeled avidin. The avidin reacts with the biotinylated antibody on the microtiter-plate.
After washing, add TMB (3, 3’, 5, 5’-tetramethylbenzidine) which is a substrate of peroxidase. After reaction, measure the absorbance of the color density of the reaction mixture at 450 nm.
In proportion to increase the collagen concentration of samples, the color density decreases since the amount of biotinyleted antibody and peroxidase labeled avidin decreases. |
<Specificity>
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Cross Reactivity
Cross reactivity comparison between human collagen type 1 (100%) and the other sources and types of collagen.
The coss reactivity of porcine collagen against human collagen is 20%. |
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Standard Curve
The concentration of collagen is calculated based on standard curve using 4-parameter formula. |
Component
| 1) |
Collagen coated microtiter plate
Pull out the strips (8 wells/strip) you wish to run from the
bag. Return the rests and store in the bag after sealed tightly
at 2~8?. Use within 30 days after unsealed. |
96 wells,
12 strips |
| 2) |
Collagen standard
Lyophilized. Reconstitute the lyophilized powder with 0.5mL
of diluent A(standard/sample diluent). The concentration of
the standard after reconstitution is printed on the label of
reagent bottle. Prior to assay, prepare a standard solution
set (see “preparation of a standard solution set”
(p.5)).
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0.5mL |
| 3) |
Biotinylated anti-collagen antibody concentrate
Ready for use. |
0.8mL |
| 4) |
Avidin-HRP conjugate concentrate
Prior to use, dilute 100 times with diluentB (see “preparation
of avidin-HRP conjugate solution” (p.6)). |
0.07mL |
| 5) |
Diluent A
Use for dilution of the collagen standards and samples. |
10mL |
| 6) |
Diluent B
Use for dilution of avidin-HRP conjugate concentrate. |
10mL |
| 7) |
Wash buffer concentrate (10X concentrated)
Dilute 10 times with distilled water or deionized water (see
preparation of wash buffer (p.6)). |
50mL |
| 8) |
TMB substrate
3, 3’, 5, 5’-tetramethylbenzidine. Ready for use. |
7mL |
| 9) |
Stop solution
1N sulfuric acid. Ready for use. |
7mL |
| 10) |
Plate seal
Cut in an appropriate size for use. |
2 sheets |
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