Dear Scientists,

I am conducting a LDH assay (roche) using A549 cell line (50000 cells/well) with complete DMEM media. I am treating the cells with different compounds with different concentration but in each concentration our result becomes negative because of higher negative control than treated ones.

The formula prescribed from the protocol was utilized;

Relative LDH release = ( X - Negative control) / (Positive control - Negative control)

Where X = sample absorbance value (@ ~ 490 nm)

In negative control, Cells treated with medium alone without compound

Positive control = 1% Triton X-100

Vehicle control = 0.5% DMSO

Blank = without cells

What would be the reason for this negative values in treatment groups?

Thanks in advance