We are facing a problem in detecting ERK-1 protein expression. We are doing knockdown experiment of ERK-1 protein. We use peptide based vehicle for siRNA delivery and it is FAM labelled which emits fluorescence at FITC channel. The siRNA we use is SignalSilence® p44/42 MAPK (Erk1/2) siRNA Catalogue number-6560, Lot No-13,Ref-09/2017 CST from CST. The primary antibody we use is p44/42 MAPK (Erk1/2) Antibody Catalogue number-9102, Lot No –23 in 1:100 dilution. And the secondary antibody we use is Alexa Fluor 568 (Cat. No. A-11036) Lot No –1924788 from Thermo Fisher Scientific in 1:500 dilution. The problem we are facing is that the secondary antibody Alexa Fluor 568 is showing much enhanced signal in the TRITC channel in the presence of the FAM labelled peptides thus making our experiment meaningless. We have also used Alexa Fluor 350 (Cat. No. A-21068), Lot No – 1904526 Thermo Fisher Scientific but did not get enough signal even in 1:100 dilution. We also have used Alexa Fluor 647 Cat no –A32728 but got very faint signal in TRITC channel. What should be done so that I get the correct expression of ERK protein in the presence of FAM labelled peptides? Waiting for your valuable suggestions.