710837

antibody from Invitrogen Antibodies

Targeting: CDC25A

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Antibody Data

Product number

710837

Provider

Invitrogen Antibodies

Product name

Phospho-Cdc25A (Ser18) Recombinant Polyclonal Antibody (20HCLC)

Antibody type

Polyclonal

Antigen

Synthetic peptide

Description

This antibody is predicted to react with Monkey and Bovine.

Antibody clone number

20HCLC

Concentration

0.5 mg/mL

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 HCT116 (Lane 1), HCT116 treated with UV (For 40 min) (Lane 2), HeLa (Lane 3), HeLa treated with UV (For 40 min) (Lane 4), HeLa treated with Nocodazole (Lane 5). The blots were probed with Anti-Cdc25a (pS18) Recombinant Rabbit Polyclonal Antibody (Product # 710837, 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 36 kDa band corresponding to Cdc25a (pS18) 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 Cdc25apS18 using Anti-Cdc25apS18 Recombinant Rabbit Polyclonal Antibody (Product # 710837, 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 Cdc25apS18 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 cytoplasmic and nuclear localization of Cdc25apS18. Panel e) represents composite image of untreated cells with no signal. Panel f) represents control cells with no primary antibody to assess background.