PeproTech-ELISA kit common problem summary

1. How many plates can PeproTech's ELISA kit (EDK) detect?

A standard ELISA kit contains enough components to detect 10 96-well plates, while a Mini ELISA kit contains enough components to detect two 96-well plates.

2. What is the stability of the HRP conjugate contained in the PeproTech ELISA kit?

The Avidin-HRP in the ABTS ELISA kit is stable for up to one month at 2-8 °C, and can be stored for up to -20 °C for up to 2 years to avoid repeated freezing and thawing. Streptavidin-HRP in the TMB ELISA kit can be stable for half a year at 2-8 °C.

3. How do I get cross-reactivity information for the kit?

PeproTech tested the cross-reactivity of its ELISA kits in the laboratory and can view the test results in the instructions.

4. What if I want to stop at the weekend during the ELISA experiment?

It can be coated with coated antibody on Friday, then stored at 4 ° C, and continue to experiment next Monday, please note: changes in antibody coating time may cause differences between the results of the plate.

5. Does PeproTech's EDK apply to all types of samples?

Although PeproTech does not test each sample with its kit, these kits should be suitable for, but not limited to, the following specimens: serum, plasma, cell culture supernatant, urine, and saliva.

6. Does the color reaction need to be terminated?

The color reaction of Avidin-HRP and substrate ABTS does not need to be terminated. When the standard zero hole OD value is less than 0.2, or the highest concentration hole OD value is less than 1.2, a reliable standard curve can usually be obtained, such as to terminate the display. For the color reaction, 1% SDS can be used as the stop solution. Although the data show that SDS is the best stop solution for ABTS, PeproTech does not use a stop solution in the laboratory.

7. Can PeproTech's ABTS EDK use TMB as a substrate?

PeproTech's EDK is optimized with ABTS, so it is best to use ABTS as a substrate. These kits are also used with TMB, but some adjustments are needed:

 Avidin-HRP in the kit cannot be paired with TMB, and streptavidin-HRP must be purchased separately;

 The working concentration of streptavidin-HRP needs to be optimized;

 When using streptavidin-HRP and TMB, it is usually necessary to use a stop solution, please follow the instructions;

 The reaction time of TMB needs to be optimized before adding stop solution;

 Use the recommended ELISA plate and read the absorbance at 450 nm with a correction wavelength of 650 nm.

8. Why do you add D-mannitol to the EDK component?

The purpose of adding D-mannose to the EDK component is to make the protein/antibody visible, which does not affect the detection of the ELISA.

9. Can I use the curve in the EDK manual as my standard curve?

Each ELISA plate must be a separate standard curve, that is, the standard curve for one plate cannot be used for the other plate. The standard provided in PeproTech's EDK instructions is made in your own laboratory and can only be used as Example.

10. How is PeproTech's standard curve generated?

When performing an ELISA in the laboratory, PeproTech reads the absorbance using a Molecular Devices microplate reader and then analyzes the results using SOFTmax PRO software, which performs a four-parameter curve fit on the acquired data.

The specific algorithm is as follows:

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