Human NANOG Transcription Factor Activity Assay by RayBiotech Inc.

Manufacturer RayBiotech Inc.  |   Model: TFEH-NANOG  |  Available Worldwide
RayBio® Human NANOG Transcription Factor Activity Assay. This assay uses a dsDNA coated plate with canonical NANOG binding sequences to semi-quantitatively detect active NANOG in lysates or nuclear extracts.


Human NANOG Transcription Factor Activity Assay by RayBiotech Inc. product image
Human NANOG Transcription Factor Activity Assay
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Introduction

Homeobox-containing transcription factor NANOG is essential in maintaining the pluripotent cells of the inner cell mass and in the derivation of embryonic stem cells (ESCs). Overexpression of NANOG is capable of maintaining the pluripotency and self-renewing characteristics of ESCs under what normally would be differentiation-inducing culture conditions. Concomitant with this essential function in pluripotent cell maintenance, expression of NANOG has restricted patterning in the inner cells of the morula, blastocyst, and epiblast during different developmental processes. Additionally, NANOG acts as a molecular gatekeeper to suppress or ensure differentiation in response to events in fluctuating environments and the presence of stimuli. NANOG has a single homeodomain that binds to DNA through the consensus sequence TAATGG. Coordinating with OCT4 and SOX2, NANOG regulates a large cohort of genes involved in pluripotency and development in later stages.

Product Features

  • Specific transcription factor-DNA binding assay
  • Perfect alternative to EMSA
  • Easy to perform in an ELISA format
  • Non-radioactive assay
  • High throughput (96-well plate format)
  • Assay can be completed within 5 hours

Specificity

The olionucleotide/antibody pair provided in this kit recognizes human NANOG in whole lysates and nuclear extracts.

Target Protein Information

Synonyms
Homeobox protein NANOG, Homeobox transcription factor Nanog (hNanog)
Accession Number
Gene Names
NANOG
Gene ID
79923

Species Detected

Human

Application Notes

Kit Components
  • 96-well Strip Microplate pre-coated with DNA probes
  • DNA Binding Buffer
  • Positive Control Sample
  • Specific Competitor DNA probe
  • Non-specific Competitor DNA probe
  • Assay Reagent
  • DTT
  • Wash Buffer
  • Primary Antibody
  • HRP-conjugated Secondary Antibody
  • TMB One-Step Substrate Reagent
  • Stop Solution
Other Materials Required
  • Distilled or deionized water
  • 100 ml and 1 liter graduated cylinders
  • Tubes to prepare sample dilutions
  • Absorbent paper
  • Precision pipettes to deliver 2 µl to 1 ml volumes
  • Adjustable 1-25 ml pipettes for reagent preparation
  • Benchtop rocker or shaker
  • Microplate reader capable of measuring absorbance at 450 nm
Protocol Outline
  1. Prepare all reagents and samples as instructed in the manual.
  2. Add 100 µl of sample or positive control to each well.
  3. Incubate 2 h at RT or O/N at 4 °C.
  4. Add 100 µl of prepared primary antibody to each well.
  5. Incubate 1 h at RT.
  6. Add 100 µl of prepared HRP-secondary antibody to each well.
  7. Incubate 1 h at RT.
  8. Add 100 µl of TMB One-Step Substrate Reagent to each well.
  9. Incubate 30 min at RT.
  10. Add 50 µl of Stop Solution to each well.
  11. Read at 450 nm immediately.

Storage/Stability

Upon receipt, the positive control should be removed and stored at -20° or -80°C. The remainder of the kit can be stored for up to 6 months at 2-8°C from the date of shipment. Opened Microplate Wells or reagents may be stored for up to 1 month at 2° to 8°C. Return unused wells to the pouch containing desiccant pack, reseal along entire edge.
Note: The kit can be used within one year if the whole kit is stored at -20°C upon receipt. Avoid repeated freeze-thaw cycles.

Product Overview

Human NANOG Transcription Factor Activity Assay by RayBiotech Inc. product image

Human NANOG Transcription Factor Activity Assay

Manufacturer RayBiotech Inc.  |  Available Worldwide

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