MA5-11355

antibody from Invitrogen Antibodies

Targeting: PCNA

Antibody data

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Validation data

Product number: MA5-11355 - Provider product page
Provider: Invitrogen Antibodies
Product name: PCNA Monoclonal Antibody (PC10), Biotin
Antibody type: Monoclonal
Antigen: Recombinant full-length protein
Description: Methanol fixation is required for immunofluorescence and flow cytometry staining. To decrease nonspecific staining for both applications, using a Biotin/Streptavidin blocker is recommended.
Conjugate: Biotin
Antibody clone number: PC10
Concentration: 200 µg/mL

PCNA protein structure - MA5-11355 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on modified whole cell extracts (1% SDS) (30 µg lysate) of HeLa (Lane 1), MCF7 (Lane 2), Jurkat (Lane 3), HEK-293 (Lane 4), NIH/3T3 (Lane 5), MDA-MB-231 (Lane 6), A549 (Lane 7), SH-SY5Y (Lane 8) and Raji (Lane 9). The blot was probed with anti-PCNA Monoclonal Antibody (Product # MA5-11355, 1:500 dilution) and detected by chemiluminescence using Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.25 µg/mL, 1:4000 dilution). A 34 kDa band corresponding to PCNA was observed across the cell lines tested.
Conjugate: Biotin

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of PCNA was achieved by transfecting HeLa cells with PCNA specific siRNAs (Silencer® select Product # s10133, s10135). Western blot analysis (Fig. a) was performed using modified whole cell extracts (1% SDS) from the PCNA knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with PCNA Monoclonal Antibody (PC10), Biotin (Product # MA5-11355, 1:1000 dilution) and Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.25µg/mL, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to PCNA.
Conjugate: Biotin

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of PCNA was performed by loading 50 µg of the indicated whole cell lysates per well, and 10 µL of PageRuler Plus Prestained Protein Ladder (Product # 26619) onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane using the G2 Fast Blotter (Product # 62288), and blocked with StartingBlock T20 (TBS) Blocking Buffer (Product # 37543) for 1 hour at room temperature. PCNA was detected between 30-35 kDa after probing with a biotinylated PCNA monoclonal antibody (Product # MA5-11355) at a dilution of 1:50 in StartingBlock T20 (TBS) Blocking Buffer (Product # 37543) overnight at 4C on a rocking platform, washing in TBST, and probing with an HRP-conjugated goat anti-mouse IgG secondary antibody (Product # 31430) at a dilution of 1:40,000 for 1 hour. Chemiluminescent detection was performed using SuperSignal West Pico (Product # 34080). Images were acquired on a Thermo Scientific myECL Imager (Product # 62236).
Conjugate: Biotin

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of PCNA (purple) in HeLa cells. Cells were fixed and permeabilized with ice-cold methanol, then blocked with 1% Blocker BSA in PBS (Product # 37525), both for 15 minutes at room temperature. Cells were stained with a biotinylated PCNA monoclonal antibody (Product # MA5-11355) at a dilution of 1:20 in 1% Blocker BSA in PBS (right panel), or incubated in blocking buffer alone as a negative control (left panel) for 1 hour at room temperature, and then incubated with Streptavidin-DyLight 550 (Product # 84542) at a dilution of 1:1000 for 1 hour at room temperature. Nuclei (blue) were stained with DAPI (Product # 46190). Images were taken on a Thermo Scientific ToxInsight Instrument at 20X magnification. NOTE: The perinuclear staining observed is due to the binding of DyLight 550-Streptavidin to endogenous biotin. To prevent such background staining, it is necessary to block the endogenous biotin.
Conjugate: Biotin

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of PCNA (purple) in HeLa cells. Cells were fixed and permeabilized with ice-cold methanol, then blocked with 1% Blocker BSA in PBS (Product # 37525) along with Solution 1 from a biotin-streptavidin blocking kit (to block endogenous biotin), both for 15 minutes at room temperature. Cells were stained with a biotinylated PCNA monoclonal antibody (Product # MA5-11355) at a dilution of 1:20 in 1% Blocker BSA in PBS containing Solution 2 from a biotin-streptavidin blocking kit (right panel), or incubated in blocking buffer alone as a negative control (left panel) for 1 hour at room temperature, and then incubated with Streptavidin-DyLight 550 (Product # 84542) at a dilution of 1:1000 for 1 hour at room temperature. Nuclei (blue) were stained with DAPI (Product # 46190). Images were taken on a Thermo Scientific ToxInsight Instrument at 20X magnification. NOTE: Blocking endogenous biotin is required to eliminate perinuclear staining.
Conjugate: Biotin