Journal: The Journal of Veterinary Medical Science
Article Title: Expression of Adrenergic and Cholinergic Receptors in Murine Renal Intercalated Cells
Figure Lengend Snippet: Characterization of M-1 cellular expression of aquaporin A) Characterization of AQP6 mRNA and protein expression in M-1 cells by RT-PCR (upper column) and immunoblot (middle column), respectively. An AQP6 specific band (arrowhead) was amplified from M-1 cellular total RNA (lane 1), M-1 cellular genomic DNA (gDNA, lane 3) and normal mouse kidney total RNA (MK cDNA, lane 4). No positive band was observed in RTase (−) negative control lane (lane 2). Immunoblot demonstrating the AQP6 specific band (arrowhead) and glycosylated band (double arrowhead) in total membrane protein extracted from M-1 cells (lane 1) and murine kidney (MK, lane 2). B) Characterization of AQP2 mRNA and protein expression in M-1 cells by RT-PCR (upper column) and immunoblot (middle column), respectively. AQP2 mRNA was not amplified from M-1 cellular total RNA (lane 1), although it was observed in M-1 gDNA (lane 3) and MK cDNA (lane 4). Similarly, AQP2 protein was not observed in total membrane protein isolated from M-1 cells (lane 1), although it was observed in total membrane protein isolated from the MK (lane 2). An arrowhead and a double arrow indicate the unglycosylated and glycosylated bands, respectively. Immunofluorescence showed the M-1 cells were positive to the AQP6 antibody (A, lower column), but negative to AQP2 antibody (B, lower column). Bar=50 µ m.
Article Snippet: Membranes were incubated with diluted primary antibodies including rabbit anti-AQP6 polyclonal antibody (pAb, #AQP-006, Alomone Labs, Jerusalem, Israel), rabbit anti-AQP2 pAb (#AQP-002, Alomone labs), rabbit anti-ADR b2 pAb (#Sc-569, Santa Cruz, TX, U.S.A.) and rabbit anti-mAChR m1 pAb (#010, Alomone Labs) for 2 hr at room temperature or at 4°C overnight.
Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Amplification, Negative Control, Isolation, Immunofluorescence