Mouse prostatic tissues were dissected from cPten−/− Luc males. After overnight fixation in 4% PFA, tissues were processed and sectioned. IHC was performed as previously described with anti-mouse AR antibody (Santa Cruz) at 1:1000 dilution (Liao et al. 2010a (link)). For IF staining, mouse NPF and CAF cells were seeded on Lab Tek II chamber slides (Thermo Scientific) with BFS medium for 24 hours, followed by fixing in 4% PFA for 10 mins and washing with PBS. After blocking, Smooth Muscle Actin (SMA) antibody conjugated with Cy3 dye (Sigma) was applied for 1 hour at a 1:60 dilution. For vimentin (Santa Cruz; 1:50 dilution) and N-Cadherin (Santa Cruz; 1:50 dilution), the primary antibody was applied for 1 hour, following by the staining of secondary antibody conjugated with either FITC or rhodamine. Samples were counter-stained with DAPI and then examined with fluorescent microscopy (Observer.Z1, ZEISS).
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Prostate Cancer Immunohistochemistry and IF
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Variable analysis
independent variables
- Mouse genotype (Pten-/- Luc males)
- Androgen receptor (AR) expression (measured by IHC)
- Smooth muscle actin (SMA) expression (measured by IF)
- Vimentin expression (measured by IF)
- N-Cadherin expression (measured by IF)
- Fixed tissue processing and sectioning procedures
- Antibody dilutions for IHC and IF staining
- Positive control: None mentioned
- Negative control: None mentioned
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