ELISA plate, can it be used instead of an ELISpot plate?

[Vsorah80]

I wonder if it ispossible to run ELISPOT assay on ELISA plate?

Thank you..

[Christian@mabtech.com]

Hello Vsorah80,

 

 

Although it is possible to run ELISpot on a normal ELISA plate (i.e you will see spots), results will be far from optimal. Both the quality and frequency of detected spots will be significantly reduced. Consequently, the sensitivity of the assay will be lowered compared to using a recommended ELISpot plate based on a PVDF membrane. 

 

The main reason for this can be found in the increased protein binding capacity of PVDF compared to plastic. You see, the quality of any ELISpot depends on secreted cytokines binding to a dense population of good capture antibody right at the vicinity of the secreting cell. After EtOH pre-treatment, PVDF is able to achieve just that due to its unique molecular properties (https://www.mabtech.com/knowledge-center/tutorials-and-guidelines/plate-coating-guide/why-use-ethanol-elispot). Spots will be sharp, optimal in terms of number and will not dissolve out into the supernatant. 

 

By contrast, an ELISA plate is not able to achieve the necessary antibody density and the resulting spots will be considerably lackluster. It does not matter how much capture antibody you add in each well, plastic plates will never get you the results that you want.

 

 

To demonstrate this difference, the attached powerpoint file below contains an image of an IL-17A ELISpot done in an ELISA plate and how this compares to our normal protocol using EtOH activated PVDF membrane. Now these results are derived from two different donors so it is impossible to compare spot-numbers. Nonetheless, the massive degradation in quality should be obvious to everyone. Please, stay away from using ELISA plates in ELISpot! 

 

ELISpot_elisaplate_vs_PVDF.pptx

 

 

 

best regards,

Christian