Overview

  • Product name

    SREBP-1 Transcription Factor Assay Kit
  • Detection method

    Colorimetric
  • Sample type

    Cell culture extracts, Nuclear Extracts
  • Assay type

    Semi-quantitative
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Product overview

    Abcam's SREBP-1 Transcription Factor Assay (ab133125) is a non-radioactive, sensitive method for detecting specific transcription factor DNA binding activity in nuclear extracts.

    A 96-well enzyme-linked immunosorbent assay (ELISA) replaces the cumbersome radioactive electrophoretic mobility shift assay (EMSA). A specific double stranded DNA (dsDNA) sequence containing the SREBP response element is immobilized onto the wells of a 96-well plate. SREBP-1 contained in a nuclear extract binds specifically to the SREBP response element. SREBP-1 is detected by addition of a specific primary antibody directed against SREBP-1. A secondary antibody conjugated to HRP is added to provide a sensitive colorimetric readout at 450 nm. The SREBP-1 Transcription Factor Assay detects human, rat, and murine SREBP-1.

  • Notes

    Lipid homeostasis in vertebrate cells is regulated by a family of transcription factors called sterol regulatory elements binding proteins (SREBPs). SREBPs directly activate the expression of over 30 gened involved in teh synthesis and uptake of cholesterol, fatty cids, triglycerides and phopholipids. SREBP-1 acts primarily to activate genes in fatty acid synthesis. Nuclear SREBP-1 upregulates gene expression of a group of a target lipogenic enzymes such as acetyl CoA carboxylase and fatty acid synthase.

  • Platform

    Microplate reader

Properties

Associated products

Images

  • HepG2 cells were grown in protein free hybridoma medium overnight (Life Technologies), followed by a 30 minutes incubation with the addition of 1% (2-Hydroxypropyl)-β-cyclodextrin (HPBC; Sigma) or with the supplement of liquid cholesterol (Life Technologies). 10 uL of nuclear extracts (ab113474) were tested, corresponding to 2.5e5 cells per well.

  • Positive control (10 or 5 uL) with (10 or 2 uL) or without inhibitor (duplicates; +/- SD).

  • Comparison of Abcam's SREBP-1 Transcription Factor Assay Kit (ab133125) and SREBP-2 Transcription Factor Assay Kit (ab133114) specificity. The results demonstrate little to no cross reactivity of SREBPs in the opposing assay. Cross Reactivity: (+) SREBP-1a, SREBP-1b, and SREBP-1c
  • Measurement of SREBP-1 activity in HeLa (squares) and HepG2 (dots) cell lysates.
  • Quantification of recombinant SREBP-1 from E. coli cell lysates.

Protocols

References

This product has been referenced in:

  • Marwarha G  et al. Palmitate-Induced SREBP1 Expression and Activation Underlies the Increased BACE 1 Activity and Amyloid Beta Genesis. Mol Neurobiol N/A:N/A (2018). Read more (PubMed: 30569418) »
  • Salomone F  et al. Silibinin Restores NAD? Levels and Induces the SIRT1/AMPK Pathway in Non-Alcoholic Fatty Liver. Nutrients 9:N/A (2017). Read more (PubMed: 28973994) »
See all 2 Publications for this product

Customer reviews and Q&As

1-2 of 2 Abreviews or Q&A

Answer

The antibody supplied in the SREBP-1 Transcription Factor Assay Kit is polyclonal. We have not mapped which portions of SREBP-1 it detects. The positive control supplied is a clarified cell lysate, so we expect it will contain a mixture of SREBP-1 forms. The antibody in this kit will detect the SREBP-1 that is bound to the immobilized DNA, so if the SREBP-1 is not able to bind to the DNA, it will not be detected in this assay.

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Answer

1. The plate is coated with the dsDNA consensus sequence specific for SREBP. The sequence is 5'TCACCTGA-3'.
2. The competitor DNA sequence is 5'TCACCTGA'.
3. The HepG2 and HeLa lysates are whole cell lysates.




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