Hi, Does anyone knows for colorimetric detection in WB, it is best to use AP or HRP conjugated? Recently, I am intend to buy the rabbit monoclonal antibody from cell signaling, but when i search for the only anti-rabbit HRP conjugated available, the information sheet describes that this HRP conjugated is design for chemiluminescence detection. isnt meaning that cannot be used for colorimetric detection?
you should be able to do that, sennsitivity can be a problem. recent HRP colored substrates are very sensitive, e.g. pierce cat#34018 (HRP) or 34042 (AP). anyway, give the cat# and lets check if this enzyme is special.
we use AP conjugated antibodies, n i find them best for colorimetric detection.. although chemiluminescence is better n has more sensitivity.. we dnt have detection facilities so colorimetric is the option.
well i also use colorimetric detection method. yes sensitivty is less n u should more protein in it.. AP is easier to do and less cumbersome compared top HRP ones if u use ready to use substrate solution of BCIP/NBT (whic i use). HRP is also fine.. we make our own substrate using DAB and H2O2.. hense seems more cumbersome.. but i find bands more clean in HRP compared to AP system..n your same HRP can be used for colorimetric detection
can HRP labelled antibody used with NBT/BCIP for colorimetric reaction in insitu hybridization on zebrafish embryos? will HRP also give color ppt as Alkaline phosphatase does?
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