Purpose and importance of the blocking step during ELISPOT a

Purpose and importance of the blocking step during ELISPOT assay?

What results would one expecte if this step had been omitted?

Solution

Enzyme –Linked immuneSPOT assay- This is a preferred assay to examine “recipient graft- rejection” at the time of organ transplantation (for example, T-cell activity & cytokine activity to antigen & antibody complexes). In this assay, polyclonal capture of antibodies onto a PVDF-backed microplate finally this plate is going to get blocked using a “non-reactive serum protein” & RPMI medium because it is going to produce “physiological PH” to the culturing cells, where it produces reducing agent glutathione & vitamins in higher proportions. Omitting this step is leading to “differential spot quality with different size & density” so that fuzzy spots may be appeared finally difficult to detect antigen binding to antibody for cell-mediated immunity