NF-kB p65 ELISA Kit

Catalog No. ABIN5526742

This Human, Rat and Mouse NF-kB p65 ELISA Kit is a Colorimetric ELISA Kit designed to quantify Human, Rat and Mouse NF-kB p65 in Adherent Cell Culture.

Quick Overview for NF-kB p65 ELISA Kit (ABIN5526742)

Target: NF-kB p65 (NFkBP65) (Nuclear Factor-kB p65 (NFkBP65))

Binding Specificity: pSer536, total

Reactivity: Human, Rat, Mouse

Detection Method: Colorimetric

Method Type: Cell ELISA

Application: ELISA

Sample Type: Adherent Cell Culture

Product Details NF-kB p65 ELISA Kit

Purpose: Cell-Based ELISA Kit. This assay semi-quantitatively measures NF-kB P65 phosphorylated at Serine-536 as well as total NF-kB P65 in adherent cell lines.

Analytical Method: Semi-Quantitative

Specificity: This ELISA kit recognizes Human, Mouse and Rat NF-kB P65 phosphorylated at site Serine-536 as well as total NF-kB P65.

Characteristics:

• Rapidly measure phosphorylated protein in adherent cell lines
• Simultaneously measure Phosphorylated protein and pan protein in one experiment (for normalization purpose)
• No sample lysis is needed
• Compatible with a standard ELISA plate reader

Components:

• Uncoated 96-well Strip Microplate
• Wash Buffers
• Fixing Solution
• Quenching Buffer
• Blocking Buffer
• Anti-phospho antibody
• Anti-pan antibody
• HRP-Conjugated Secondary Antibody
• TMB One-Step Substrate
• Stop Solution

Material not included:

• Distilled or deionized water
• 100 mL and 1 liter graduated cylinders
• Tubes to prepare sample dilutions
• Protease and Phosphatase inhibitors
• 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

Application Details

Application Notes: Optimal working dilution should be determined by the investigator.

Plate: Pre-coated

Protocol:

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 2.5 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 1X HRP-Streptavidin 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.

Restrictions: For Research Use only

Handling

Storage: -20 °C

Storage Comment: Upon receipt, the kit should be stored at -20 °C. Please use within 6 months from the date of shipment. After initial use, Wash Buffer Concentrate (Item B), Assay Diluent (Item E), TMB One-Step Substrate Reagent (Item H), HRP-Streptavidin (Item G), Stop Solution (Item I) and Cell Lysate Buffer (Item J) should be stored at 4 °C to avoid repeated freeze-thaw cycles. Return unused wells to the pouch containing desiccant pack, reseal along entire edge and store at -20 °C. Reconstituted Positive Control (Item K) should be stored at -70 °C.

Expiry Date: 6 months

Target Details for NF-kB p65

Target: NF-kB p65 (NFkBP65) (Nuclear Factor-kB p65 (NFkBP65))

Alternative Name: NF-kB P65

Gene ID: 5970

UniProt: Q04206, Q04207, Q7TQN4

Pathways: NF-kappaB Signaling, RTK Signaling, TCR Signaling, TLR Signaling, Fc-epsilon Receptor Signaling Pathway, Neurotrophin Signaling Pathway, Activation of Innate immune Response, Cellular Response to Molecule of Bacterial Origin, Hepatitis C, Toll-Like Receptors Cascades, S100 Proteins