MICA ELISA Kit

Produktnummer ABIN8070042

Dieses Colorimetric ELISA kit dient der quantitativen Messung von Human MICA.

Kurzübersicht für MICA ELISA Kit (ABIN8070042)

Target: MICA (MHC Class I Polypeptide-Related Sequence A (MICA))

Bindungsspezifität: S-acylated, total

Reaktivität: Human

Nachweismethode: Colorimetric

Methodentyp: Sandwich ELISA

Applikation: ELISA

Proben: Cell Lysate, Tissue Lysate

Produktdetails MICA ELISA Kit

Verwendungszweck: Measures s-acylated MICA and total MICA in human cell and tissue lysates, enabling precise assessment of MICA s-acylation by ELISA. This assay can be developed for other species (mouse, rat, etc.) upon request. Contact techsupport@raybiotech.com.

Analytische Methode: Quantitative

Bestandteile:

Size / Qty
• Microplate Coated With Anti-Pan-MICA Antibody
• Positive Control
• HRP-Conjugated Streptavidin
• 200 μL
• Wash Buffer
• Assay Diluent
• Lysis Buffer
• TMB One-Step Substrate Reagent
• Stop Solution
• S-Acylation Buffer A
• S-Acylation Buffer B
• S-Acylation Buffer C
• S-Acylation Buffer D
• S-Acylation Blocking Reagent
• S-Acylation Reduction Reagent
• S-Acylation Labeling Reagent

Benötigtes Material:

• Acetone (pre-chilled (-20 °C))
• Microplate reader capable of measuring absorbance at 450 nm
• Protease and Phosphatase inhibitors
• Precision pipettes to deliver 2 μL to 1 mL volumes
• Adjustable 1-25 mL pipettes for reagent preparation
• 100 mL and 1 liter graduated cylinders
• Log-log graph paper or computer and software for ELISA data analysis
• Absorbent paper
• Tubes to prepare positive control or sample dilutions
• Distilled or deionized water
• Shaker

Anwendungsinformationen

Applikationshinweise: Optimal working dilution should be determined by the investigator.

Plattentyp: Pre-coated

Protokoll:

1. Prepare all reagents, samples and Positive Control as instructed.
2. Block unmodified free cysteines using prepared S-Acylation Blocking Buffer at 50 °C with gentle rocking.
3. Precipitate proteins with ice-cold acetone.
4. Wash protein pellet twice with cold acetone/water mixture.
5. Resuspend protein pellets in Buffers and adding Reducing and Labeling Reagents.
6. Precipitate proteins with ice-cold acetone.
7. Resuspend protein pellets.
8. Add 100 μL sample and Positive Control to each well. Incubate 2.5 hours at room temperature or overnight at 4 °C with gentle shaking.
9. Add 100 μL prepared HRP-Conjugated Streptavidin solution. Incubate for 1 hour at room temperature with gentle shaking.
10. Add 100 μL TMB One-Step Substrate Reagent to each well. Incubate 30 minutes at room temperature.
11. Add 50 μL Stop Solution to each well. Read at 450 nm immediately.

Beschränkungen: Nur für Forschungszwecke einsetzbar

Handhabung

Lagerung: -20 °C

Informationen zur Lagerung: The entire kit may be stored at -20°C for up to 6 months from the date of shipment. Avoid repeated freeze-thaw cycles.

Details zu MICA

Target: MICA (MHC Class I Polypeptide-Related Sequence A (MICA))

Andere Bezeichnung: MICA

Hintergrund:

Gene Symbols: MICA, PERB11.1

Background: The S-Acylated MICA ELISA Kit uses a modified "biotin-switch" method to allow for the direct visualization of human S-Acylated MICA by ELISA. In this S-Acylation biotin switch assay, unmodified free cysteines are first blocked. S-Acylated cysteines are then selectively reduced for specific labeling with biotin-maleimide reagents, which irreversibly bind to the cysteine thiol that was S-Acylated. Biotinylation of the newly formed thiol groups can then be detected by ELISA. An anti-pan MICA antibody has been coated onto a 96-well plate. Biotinylated samples are pipetted into the wells, and MICA present in a sample is bound to the wells by the immobilized antibody. The wells are washed, and HRP-conjugated streptavidin is used to detect S-Acylated MICA. The wells are again washed, a TMB substrate solution is added to the wells and color develops in proportion to the amount of S-Acylated MICA bound. The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm.

Protein Name (Synonyms): MHC class I polypeptide-related sequence A (MIC-A)

Gen-ID: 100507436

UniProt: Q29983

Pathways: Activation of Innate immune Response, Transition Metal Ion Homeostasis, Human Leukocyte Antigen (HLA) in Adaptive Immune Response