PARP-1 antibody | knockout validation | Serotec MCA1522G

This is a knockout-validated antibody summary, based on the publication "Overlapping roles for PARP1 and PARP2 in the recruitment of endogenous XRCC1 and PNKP into oxidized chromatin", as cited below [1]. Labome curates formal publications to compile a list of antibodies with unambiguous specificity within Validated Antibody Database (VAD).

PARP-1 antibody | knockout validation | Serotec MCA1522G figure 1
Figure 1. Wild type (WT), PARP1-/- , PARP2-/- , PARP3-/- and XRCC1-/- RPE-1 clonal cell lines were analysed for loss of the targeted protein by (A) western blotting and (B) immunofluorescence. From [1].
Antibody information

Mouse monoclonal IgG1

Company: Serotec

Antibody: PARP-1

Catalog number: MCA1522G

Summary: Mouse monoclonal IgG1 against human PARP-1. Reacts with human, hamster, mouse, Drosophila, Xenopus and rat. Suitable for western blot, immunofluorescence, immunohistochemistry (paraffin and frozen), immunoprecipitation, and ELISA.

Validation Method

Western blot | Immunocytochemistry

Sample

Wild type (WT) and PARP1-/- RPE-1 clonal cell lines. IC: Cells grown on coverslips were washed with PBS and fixed with 4% paraformaldehyde in PBS 10 min at RT.

Blocking agent

IC: 10% FCS in PBS for at least 30 min.

Primary incubation

IC: 60 min, RT

Secondary incubation

WB: HRP-conjugated goat anti-mouse (Bio-Rad; 170- 6516).

IC: goat anti-mouse Alexa 488 (Invitrogen; A11001), goat anti-mouse Alexa 555 (Invitrogen; A21422) or donkey anti-mouse Alexa 647 (Invitrogen; A39571) for 60 min, RT.

Detection

WB: ECL reagent (GE Healthcare) and Amersham Hyperfilm ECL (GE Healthcare) or ImageQuant LAS-4000 system connected with high-sensitivity Super CCD camera (GE Healthcare).

IC: High-resolution microscopy of fixed samples was carried out on a Zeiss AxioObserver.Z1 microscope, equipped with oil immersion objectives (PlanApochromat 63x/1.4), Hamamatsu ORCA-Flash4.0 LT camera and ZEN 2 core imaging software.

Notes

This antibody is also validated by IC in the same Figure.

Rabbit polyclonals anti-XRCC1 (Millipore; ABC738), anti-PARP2 (Active Motif; 39743) are also KO validated (by WB and IC).

Clone note

The same clone (A6.4.12) is sold as Bio-Rad MCA1522G; LifeSpan Biosciences LS-B3432; Abnova MAB5193.

References
  1. Hanzlikova H, Gittens W, Krejcikova K, Zeng Z, Caldecott K. Overlapping roles for PARP1 and PARP2 in the recruitment of endogenous XRCC1 and PNKP into oxidized chromatin. Nucleic Acids Res. 2016;: pubmed