701720

antibody from Invitrogen Antibodies

Targeting: CHEK2 bA444G7, CDS1, CHK2, HuCds1, PP1425, RAD53

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Flow cytometry (Supportive data in Antibodypedia)

Antibody Data

Product number

701720

Provider

Invitrogen Antibodies

Product name

Phospho-CHK2 (Thr387) Recombinant Rabbit Monoclonal Antibody (45H21L9)

Antibody type

Monoclonal

Antigen

Synthetic peptide

Reactivity

Human

Host

Rabbit

Isotype

IgG

Antibody clone number

45H21L9

Vial size

100 µg

Concentration

0.5 mg/mL

Storage

Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis was performed on whole cell extracts (30 µg lysate) of COLO 205 (Lane 1), COLO 205 treated with UV (For 40 min) (Lane 2), COLO 205 treated with Doxorubicin (500 nM for 3 hours) (Lane 3), A549 (Lane 4), A549 treated with UV (For 40 min) (Lane 5), A549 treated Doxorubicin (500 nM for 3 hours) (Lane 6). The blots were probed with Anti-CHK2 (pT387) Recombinant Rabbit Monoclonal Antibody (Product # 701720, 1-2 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). A 62 kDa band corresponding to CHK2 (pT387) was observed according to the treatment. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 10% Bis-Tris gel (Product # NP0301BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescence was performed on fixed and permeabilized HeLa cells treated with UV (302nM) for 60 minutes for detection of CHK2 (pT387) using Anti-CHK2 (pT387) Recombinant Rabbit Monoclonal Antibody (Product # 701720, 2 µg/mL) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 1:2000). Panel a) shows representative cells that were stained for detection and localization of CHK2 (pT387) protein (green), Panel b) is stained for nuclei (blue) using SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). Panel c) represents cytoskeletal F-actin staining using Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d) is a composite image of Panels a, b and c clearly demonstrating nuclear localization of CHK2 (pT387). Panel e) represents merged image of untreated cells with no signal Panel f) represents control cells with no primary antibody to assess background.

Flow cytometry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Flow Cytometry analysis of CHK2 [pT387] was performed on HeLa cells treated with UV (30 minutes), labeled with ABfinity™ Anti-CHK2 [pT387] Recombinant Rabbit Monoclonal Antibody (Product# 701720, 2-4 ug/ 1M cells) or with Rabbit isotype control and detected with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, (Alexa Fluor® 488 conjugate, Product # A27034, 0.4 ug/ml, 1:2500) as represented by the red and pink histograms respectively. The purple histogram represents unstained control cells and the green histogram represents no-primary-Antibody control. A representative of 10,000 cells were acquired and analyzed for each sample using an Attune® Acoustic Focusing Cytometer (4468770).