Mouse PSP94 expression is prostate tissue-specific as demonstrated by a comparison of multiple antibodies against recombinant proteins.
Prostate tissue-specific gene expression is crucial for driving potentially therapeutic genes to target specifically to the prostate. Prostate secretory protein of 94 amino acids (PSP94), also known as beta-MSP (microseminoprotein), is one of the three most abundant secretory proteins of the prostate gland, and is generally considered to be prostate tissue-specific. We have previously demonstrated that the expression of the rat PSP94 gene is strictly prostate tissue-specific by an antibody against a recombinant rat PSP94. In order to study prostate targeting utilizing the PSP94 gene in a mouse pre-clinical experimental model, we need to establish antibodies against mouse PSP94 to confirm if it is prostate tissue-specific as well. In this study, firstly we raised a polyclonal antibody against a recombinant glutathione-S-transferase- (GST-) mouse mature form of PSP94. However, it showed very poor immunoreactivity against prostate tissue PSP94 as tested in Western blotting experiments. Neither antibodies against rat PSP94 nor mouse PSP94 showed significant cross-reactivity. Thus a second antibody was established against a recombinant mouse mature PSP94 containing N-terminal polyhistidines, and stronger immunoreactivity against mouse prostate tissue PSP94 was identified in Western blotting experiments. Both of these antibodies showed immunohistochemical reactivity, while the latter showed stronger reactivity in IHC when tested with different fixatives. By studying tissue distribution, we demonstrated that, as with rat PSP94, mouse PSP94 is strictly prostate tissue-specific in experiments of both Western blotting and immunohistochemistry (IHC). This conclusion was also derived from a comparison among antibodies against human, rat, and mouse PSP94, showing very different immunoreactivities in Western blotting and IHC. Finally, a competitive assay between different species was performed. We demonstrated that antibodies against PSP94 from different species (human, primate, rodents) have poor cross-reactivities. These observations also indicate that the PSP94 gene is a rapidly evolving gene in all species. Results from this study have led to the possibility of utilizing PSP94 as a targeting agent specifically to the prostate in a mouse experimental model.
Thota
Anuradha
A
Department of Surgery, University of Western Ontario, London, Ontario, N6A 4G5 Canada.
Karajgikar
Meera
M
Duan
Wenming
W
Gabril
Manal Y
MY
Chan
Franky L
FL
Wong
Yong-Chuan
YC
Sakai
Hideki
H
Chin
Joseph L
JL
Moussa
Madeleine
M
Xuan
Jim W
JW
D000818:Animals; D000906:Antibodies / Q000276:immunology; D015153:Blotting, Western; D003429:Cross Reactions; D004195:Disease Models, Animal; D005404:Fixatives; D015870:Gene Expression; D015693:Gene Products, nef; D015316:Genetic Therapy; D005822:Genetic Vectors; D005982:Glutathione Transferase; D007150:Immunohistochemistry; D008297:Male; D051379:Mice; D011467:Prostate / Q000378:metabolism*; D011471:Prostatic Neoplasms / Q000235:genetics / Q000628:therapy; D029584:Prostatic Secretory Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology
D000906:Antibodies; D005404:Fixatives; C091445:GST-Nef protein, recombinant; D015693:Gene Products, nef; D029584:Prostatic Secretory Proteins; D011993:Recombinant Fusion Proteins; C043076:beta-microseminoprotein; D005982:Glutathione Transferase