Uromucoid Polyclonal Antibody

WB, IHC-P, IHC-F, IF

22242

Purified Rabbit Polyclonal Antibody (Pab)

Tissue/cell: mouse kidney tissue; 4% Paraformaldehyde-fixed and paraffin-embedded;
Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Block endogenous peroxidase by 3% Hydrogen peroxide for 30min; Blocking buffer (normal goat serum, C-0005) at 37℃ for 20 min;
Incubation: Anti-MCKD2/UMOD Polyclonal Antibody, Unconjugated(bs-2189R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining

Sample:
Lane1: Kidney(Rat) Lysate at 30 ug
Lane2: Kidney carcinoma(Human) Lysate at 30 ug
Primary: Anti-MCKD2/UMOD (bs-2189R) at 1:200 dilution;
Secondary: HRP conjugated Goat Anti-Rabbit IgG(bs-0295G-HRP) at 1: 3000 dilution;
Predicted band size : 65kD
Observed band size : 65kD

Sample: Kidney(Mouse) lysate at 30 ug;
Primary: Anti-Uromucoid (bs-2189R) at 1:300 dilution;
Secondary: HRP conjugated Goat-Anti-rabbit IgG(bs-0295G-HRP) at 1: 5000 dilution;
Predicted band size:61/65 kD
Observed band size:65 kD

Tissue/cell: rat kidney tissue; 4% Paraformaldehyde-fixed and paraffin-embedded;
Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Blocking buffer (normal goat serum, C-0005) at 37℃ for 20 min;
Incubation: Anti-Uromucoid Polyclonal Antibody, Unconjugated(bs-2189R) 1:200, overnight at 4°C; The secondary antibody was Goat Anti-Rabbit IgG, Cy3 conjugated(bs-0295G-Cy3)used at 1:200 dilution for 40 minutes at 37°C. DAPI(5ug/ml, blue, C-0033) was used to stain the cell nuclei

Tissue/cell: human kidney tissue; 4% Paraformaldehyde-fixed and paraffin-embedded;
Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Blocking buffer (normal goat serum, C-0005) at 37℃ for 20 min;
Incubation: Anti-Uromucoid Polyclonal Antibody, Unconjugated(bs-2189R) 1:200, overnight at 4°C; The secondary antibody was Goat Anti-Rabbit IgG, Cy3 conjugated(bs-0295G-FITC)used at 1:200 dilution for 40 minutes at 37°C. DAPI(5ug/ml, blue, C-0033) was used to stain the cell nuclei

Blank control: Mouse kidney (blue).
Primary Antibody:Rabbit Anti-Coxsackie Adenovirus Receptor antibody (bs-2189R, Green); Dilution: 1 µg in 100 µL 1X PBS containing 0.5% BSA;
Isotype Control Antibody: Rabbit IgG(orange) , used under the same conditions;
Secondary Antibody: Goat anti-rabbit IgG-FITC(white blue), Dilution: 1:200 in 1 X PBS containing 0.5% BSA.
Protocol
The cells were fixed with 2% paraformaldehyde for 10 min at 37℃. Primary antibody (bs-2189R, 1 µg /1x10^6 cells) were incubated for 30 min at room temperature, followed by 1 X PBS containing 0.5% BSA + 10% goat serum (1 hour) to block non-specific protein-protein interactions. Then the Goat Anti-rabbit IgG/FITC antibody was added into the blocking buffer mentioned above to react with the primary antibody at 1/200 dilution for 40 min at room temperature. Acquisition of 20, 000 events was performed.