I assume you mean colourimetric assay, how do you do the TRAP assay?

TRAP staining and quantifying

Aspirate medium and wash cells with 200µl PBS. Aspirate PBS.

Add 100µl Citrate buffer + 0.1% Triton X-100, incubate 5 minutes @RT.

Add 100µl pNPP solution, incubate 5 minutes @RT.

Stop reaction by adding 50µl 0.5M NaOH.

Read OD at 405nm.

pNPP solution:

pNPP-40mg (1 capsule)

Tartrate solution-1.8ml

Citrate buffer-5.7ml

I found in the past that making the substrate up from the original pNP and grinding it up in DMF and the diluting it in aqueous buffer if the powder was not dissolved well enough the background was grainy.  You could try centrifuging or filtering the pNPP and do the reaction at 37C.

All the best

I also find that TRAP stain works best when all ingredients are made fresh right before use and the reaction is carried out at 37C. Also consider your reaction times. They may need some adjustment.

Did you know your microplate reader filters has an maximum and optimal wavelength? Because it is important when you have low amount of cells and weak enzymes activity. Filters that differ from the optimal wavelength more than +/- 20 nm could affect detecting the absorbance and or fluorescence emission of dyes and reagents,  the further they are from the peak the weaker the signal will be, which can lead to higher background and noise.