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Instruction manual for human cytochrome C (CytC) ELISA kit
Detection range: 96T Purpose of use: Experimental principle kit composition | 1 | 30x concentrated wash solution | 20ml × 1 bottle | 7 | Termination solution | 6 ml × 1 bottle |
| 2 | Microplate reagents | 6 ml × 1 bottle | 8 | Standard (400 nmol/L) | 0. 5 ml × 1 bottle |
| 3 | Microplate-coated plates | 12 holes × 8 | 9 | Standard dilution | 1. 5 ml × 1 bottle |
| 4 | Sample dilution | 6 ml × 1 bottle | 10 | Instructions | 1 serving |
| 5 | Color developer A liquid | 6 ml × 1 bottle | 11 | Sealing film | 2 sheets |
| 6 | Color developer B solution | 6ml × 1/bottle | 12 | Sealed bags | 1 pcs |
Specimens require procedures
- Dilution of standards: This kit provides a standard in the original times, and the user can dilute
it in a small test tube according to the following diagram.
| 200 nmol/L | Standard No. 5 |
150 μl of the original standard is added to 150 μl of the standard dilution |
| 100 nmol/L | Standard No. 4 |
150 μl of Standard 5 is added to 150 μl of standard dilution |
| 50 nmol/L | Standard No. 3 |
150 μl of Standard 4 was added to 150 μl of standard dilution |
| 25 nmol/L | Standard No. 2 |
150 μl of Standard 3 was added to 150 μl of standard dilution |
| 12. 5 nmol/L |
Standard No. 1 |
150 μl of Standard 2 was added to 150 μl of standard dilution |
- Dosing: Blank wells are set up separately (blank control wells do not add samples and microplate reagents, and the other steps are the same), standard wells, and sample wells
to be measured.
Accurately add 50 μl of the standard on the microplate-coated plate, first add 40 μl of the sample dilution to the well of the sample to be measured, and then add 10 μl of the sample to be measured (the final dilution of the sample is 5 times).
Add the sample to the bottom of the microplate plate well, trying not to touch the well wall, and gently shake the mixture
. - Incubation: Incubate at 37 °C after sealing the plate with sealing membrane for 30 min
.
- Dosing: Dilute 30x concentrated wash solution with distilled water 30x and set aside
- Washing: Carefully remove the sealing film, discard the liquid, shake dry, fill each well with washing solution, let stand for 30 seconds and discard, repeat 5 times, pat dry
. - Add enzymes: Add 50 μl of microplate reagent per well, except for blank wells
. - Incubation: Operation same as 3
. - Washing: Operation with 5
. - Color rendering: First add color developer A50μl to each well, then add color developer B50μl, gently shake and mix well, 37 °C avoid light for 10 minutes.
- Stop: Add 50 μl of stop solution per well to terminate the reaction (at this point the blue vertical to yellow)
is added. - Assay: Zero out the blank wells, and measure the absorbance (OD value)
of each well sequentially at a wavelength of 450 nm.
The assay should be performed
within 15 min after adding the stop solution.
