RAF1 ELISA Kit

Catalog No. ABIN2748581

Human, Mouse and Rat RAF1 ELISA Kit Colorimetric assay for quantification of Human, Mouse and Rat RAF1.

Quick Overview for RAF1 ELISA Kit (ABIN2748581)

Target: RAF1 (V-Raf-1 Murine Leukemia Viral Oncogene Homolog 1 (RAF1))

Binding Specificity: pSer301, total

Reactivity: Human, Mouse, Rat

Detection Method: Colorimetric

Method Type: Sandwich ELISA

Application: ELISA

Sample Type: Cell Lysate, Tissue Lysate

Product Details RAF1 ELISA Kit

Purpose: Human/Mouse/Rat Phospho-Raf-1 (Ser301) and Total Raf-1 ELISA Kit. This assay semi-quantitatively measures phophorylated Raf-1 (Ser301) and Total Raf-1 in lysate samples.

Analytical Method: Semi-Quantitative

Specificity: The antibody pair provided in this kit recognizes human, mouse, and rat Raf-1 phosphorylated at site Serine-301 and Total Raf-1.

Characteristics:

• Simultaneously measure Phosphorylated protein and pan protein in one experiment (for normalization purpose)
• Screen numerous different cell lysates without performing a Western Blot analysis
• Minimal hands-on time, convenient, and non-radioactive material

Components:

• Pre-Coated 96-well Strip Microplate
• Wash Buffer
• Anti-Phospho Antibody
• Anti-Pan Antibody
• HRP-Conjugated Secondary Antibody
• Streptavidin-Conjugated HRP
• Assay Diluent
• TMB One-Step Substrate
• Stop Solution
• Lysis Buffer
• Positive Control Sample

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

Sample Volume: 100 μL

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.

Assay Procedure:

Prepare all reagents and samples as instructed in the manual.
Add 100 μL of sample or positive control to each well.
Incubate 2.5 h at RT or O/N at 4 °C.
Add 100 μL of prepared primary antibody to each well.
Incubate 1 h at RT.
Add 100 μL of prepared 1X HRP-Streptavidin to each well.
Incubate 1 h at RT.
Add 100 μL of TMB One-Step Substrate Reagent to each well.
Incubate 30 min at RT.
Add 50 μL of Stop Solution to each well.
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 RAF1

Target: RAF1 (V-Raf-1 Murine Leukemia Viral Oncogene Homolog 1 (RAF1))

Alternative Name: Raf-1

Background: RAF proto-oncogene serine/threonine-protein kinase (Raf-1) phosphorylated at Serine-301 and Total 301

Gene ID: 5894

UniProt: P04049

Pathways: MAPK Signaling, RTK Signaling, Fc-epsilon Receptor Signaling Pathway, Neurotrophin Signaling Pathway, cAMP Metabolic Process, Stem Cell Maintenance, Hepatitis C, Autophagy, Signaling of Hepatocyte Growth Factor Receptor, VEGF Signaling, BCR Signaling