CREB1 antibody | knockout validation | Cell Signaling 9197
DOI
//dx.doi.org/10.13070/ko.en.7.1984
Date
2017-01-20

This is a knockout-validated antibody summary, based on the publication "Wnt/β-catenin pathway transactivates microRNA-150 that promotes EMT of colorectal cancer cells by suppressing CREB signaling" [1] and "Supernova: A Versatile Vector System for Single-Cell Labeling and Gene Function Studies in vivo" [2]. Labome curates formal publications to compile a list of antibodies with unambiguous specificity within Validated Antibody Database (VAD).

CREB1 antibody | knockout validation | Cell Signaling 9197 figure 1
Figure 1. Western blotting for CREB1 expression in HCT116 cells after knockout by CRISPR/Cas9. GAPDH was used as the loading control. From [1].
Antibody information

Rabbit monoclonal IgG

Company: Cell Signaling

Antibody: CREB1

Catalog number: 9197

Summary: Rabbit monoclonal IgG against a recombinant protein specific to the amino terminus of human CREB-1 protein. Reacts with human, mouse, rat, monkey, and D. melanogaster. Suitable for western blot, immunoprecipitation, immunohistochemistry (paraffin and frozen), immunofluorescence/immunofluorescence (frozen), flow cytometry and ChIP.

Article "Wnt/β-catenin pathway transactivates microRNA-150 that promotes EMT of colorectal cancer cells by suppressing CREB signaling"
Validation Method

Western blot

CREB1 antibody | knockout validation | Cell Signaling 9197 figure 2
Figure 2. Hippocampal CA1 region of wild-type mice electroporated with Supernova-CRISPR/Cas9 control (without targeting sequence in sgRNA) and Supernova-CRISPR/Cas9 targeting mouse Creb1 (SnCRISPR-Creb1). From [2].
Sample

Control and CREB1-knockout HCT116 cells. Total protein was extracted using TRIzol reagent (Invitrogen) and precipitate was lysed in the buffer containing 1% DTT, 4% CHAPS, 7 M urea, 2 M thiourea and 2% ampholine.

Article "Supernova: A Versatile Vector System for Single-Cell Labeling and Gene Function Studies in vivo"
Validation Method

Immunohistochemistry

Sample

Hippocampal CA1 region of wild-type control mice and Supernova-CRISPR/Cas9 targeting mouse Creb1. Brains were fixed in 4%PFA and cut on a freezing microtome (ROM-380; Yamato) into coronal sections at 100 μm.

Blocking agent

5% normal goat or donkey serum (Sigma) in 0.2% Triton X-100, 0.1 M PB for 1 hour at room temperature.

Primary incubation

1:1,000 dilution at 4 °C for overnight.

Secondary incubation

1:1,000 dilution Alexa 568 goat anti-Rabbit IgG(H+L) (Invitrogen) at 4 °C for overnight.

Detection

Fluorescent images were acquired using a TCS SP5 confocal microscope (Leica).

Clone note

The same clone (48H2) is sold as Cell Signaling Technology 9197, 4034S; Sigma-Aldrich C8977.

References
  1. Guo Y, Wang L, Li B, Xu H, Yang J, Zheng L, et al. Wnt/β-catenin pathway transactivates microRNA-150 that promotes EMT of colorectal cancer cells by suppressing CREB signaling. Oncotarget. 2016;: pubmed publisher
  2. Luo W, Mizuno H, Iwata R, Nakazawa S, Yasuda K, Itohara S, et al. Supernova: A Versatile Vector System for Single-Cell Labeling and Gene Function Studies in vivo. Sci Rep. 2016;6:35747 pubmed publisher