Per RNA C.C. They are reporting that Immunofl. [IMF], the method SRPT used to quantify dystrophin in the "best" method. Not Western blot, not PCR. it sounds like Prosensa has already been informed that Glaxo will sever contract.
Sentiment: Strong Buy
I have done all 3 assays with my own hands, tons of them. If the antibody used in the assay is good, and the quantification is designed and carried out properly, immunofluorescence is the best among the 3, head and shoulder. But when the antibody is not good enough, western blot is better but requires a larger amount of sample and does not tell where inside a cell or even which cell the protein in question is present. As to PCR, it is different as it does not measure protein.
The best approach is to use all 3 methods. immunofluorescence requires the sample to be fresh (never been frozen). In case of muscle sample from a human being (instead of from a tiny worm or fly), there is NO REASON for SRPT or RNA not to use all 3 assays on EVERY patients.
Umm Fl does not require sample to be frozen, it is done on fixed paragon embedded tissues all the time, but this depends on antibody reacting with structure that does not denature with fixing tissue, not all ag/ab combos work. Intracellular antigens like dystrophin, require permeablization. Westerns are nice and easier, but nothing on localization. The strength of IF here is the antibody used and ability to see localization into proper intracellular complex.
And using all 3 assays is what FDA is asking SRPT to do for the 4th biopsy, IMO. FDA should have specified where to take the sample, and all 3 assays done on all samples already, if the scientists at FDA were trained properly.
“Per RNA C.C. They are reporting that Immunofl. [IMF], the method SRPT used to quantify dystrophin in the "best" method. Not Western blot, not PCR. it sounds like Prosensa has already been informed that Glaxo will sever contract.”
Western Blot and immunofluorescence assays both detect protein using antibodies. The read-out and sample material are different but many of their properties are similar; sensitivity and semi-quantifiable.
PCR is another animal and can be both highly quantifiable and sensitive.
There is NO one “best” assay. They all have strengths and weaknesses.