ADAM9 Mouse mAb: Protocol & Application Guide

ADAM9 Mouse mAb is a mouse-derived monoclonal antibody specifically targeting human ADAM9 protein, formulated in PBS and designed for reliable detection of ADAM9 in a range of immunological applications.

Product SKU

Ab351140

Clone Source

Mouse monoclonal

Formulation

PBS (Phosphate-Buffered Saline)

Target Antigen

ADAM9 (A Disintegrin And Metalloproteinase 9)

Supplier Reference

absea #KC-8411

Recommended Storage

Store at 4°C for short-term; -20°C for long-term; avoid repeated freeze-thaw cycles

Background: ADAM9 Biology and Antibody Overview

ADAM9 (A Disintegrin And Metalloproteinase 9), also known as MDC9 or Meltrin-γ, is a member of the ADAM family of transmembrane and secreted zinc-dependent metalloproteinases. ADAM9 participates in ectodomain shedding of membrane-anchored proteins, cell adhesion, and cell–matrix interactions. It is implicated in tumour invasion, angiogenesis, and tissue remodelling, making it a target of interest in oncology and cell biology research.

The ADAM9 Mouse mAb (Ab351140, absea #KC-8411) is a mouse-derived IgG monoclonal antibody raised against human ADAM9. Formulated in PBS, it offers consistent lot-to-lot specificity and is suitable for researchers investigating ADAM9 expression and function in human cell lines and tissues.

Sample Preparation and Recommended Protocol

Cell and Tissue Lysate Preparation (Western Blot):

1. Harvest cells by scraping or trypsinisation on ice. Wash twice with ice-cold PBS.
2. Lyse in RIPA buffer supplemented with protease inhibitor cocktail and 1 mM PMSF for 30 min on ice.
3. Centrifuge at 12,000 × g for 15 min at 4°C. Collect the supernatant.
4. Determine protein concentration by BCA or Bradford assay. Load 20–40 µg total protein per lane.
5. Resolve by SDS-PAGE (8–12% polyacrylamide gel) and transfer to PVDF or nitrocellulose membrane.
6. Block membrane with 5% non-fat dry milk in TBST for 1 hour at room temperature.
7. Incubate with ADAM9 Mouse mAb at the recommended working dilution (see key facts; confirm with datasheet) overnight at 4°C.
8. Wash 3 × 10 min with TBST, then incubate with appropriate HRP-conjugated anti-mouse secondary antibody for 1 hour at room temperature.
9. Wash 3 × 10 min, then detect with ECL substrate.

Tissue Section Preparation (IHC-P):

1. Fix tissues in 10% neutral-buffered formalin and embed in paraffin. Cut 4–6 µm sections.
2. Dewax and rehydrate through graded alcohols to water.
3. Perform antigen retrieval in 10 mM citrate buffer pH 6.0 using a pressure cooker or microwave for 10–20 min.
4. Block endogenous peroxidase with 3% H₂O₂ for 10 min, then block with 5% normal serum for 30 min.
5. Apply ADAM9 Mouse mAb at the appropriate dilution; incubate overnight at 4°C in a humidified chamber.
6. Apply secondary antibody and detection reagent per the IHC kit instructions. Counterstain with haematoxylin. Mount and image.

Storage, Handling, and Quality Notes

Storage Conditions: The ADAM9 Mouse mAb (Ab351140) is formulated in PBS. For short-term use (up to 1 month), store at 4°C. For long-term storage, aliquot and store at -20°C. Avoid repeated freeze-thaw cycles, which can denature the antibody and reduce activity. Do not store in frost-free freezers.

Handling: Centrifuge the vial briefly before opening to collect contents. Work on ice when preparing dilutions. Use sterile, low-protein-binding tubes to minimise loss due to adsorption.

Quality: As a mouse monoclonal antibody, ADAM9 Mouse mAb offers high reproducibility and epitope specificity. Include positive and negative controls in every experiment. Confirmed positive control cell lines or tissues expressing ADAM9 should be used to validate the assay before testing unknown samples.

Troubleshooting

Use the table below to diagnose and resolve common issues encountered when using ADAM9 Mouse mAb:

Problem	Possible Cause	Suggested Solution
No signal on Western blot	Antibody dilution too high; insufficient antigen; incomplete transfer	Reduce antibody dilution; increase protein loading; verify transfer with Ponceau S staining
High background	Insufficient blocking; secondary antibody cross-reactivity; antibody concentration too high	Extend blocking time or use 5% BSA; titrate primary antibody; include additional TBST washes
Multiple non-specific bands	Protease degradation; antibody cross-reactivity	Ensure fresh protease inhibitors in lysis buffer; confirm band size against expected ADAM9 MW
Weak IHC signal	Insufficient antigen retrieval; antibody dilution too high	Optimise antigen retrieval time/method; reduce working dilution of antibody
Loss of antibody activity	Repeated freeze-thaw; improper storage	Aliquot upon receipt; store at -20°C; avoid frost-free freezers

Frequently asked questions

What applications is ADAM9 Mouse mAb (Ab351140) validated for?

The product datasheet should be consulted for the complete and up-to-date list of validated applications. Typical applications for antibodies of this class include Western blotting (WB), immunohistochemistry on paraffin sections (IHC-P), and immunofluorescence (IF). Please confirm the specific validated applications with Aladdin Scientific customer support or the product datasheet.

What is the recommended storage condition for ADAM9 Mouse mAb?

ADAM9 Mouse mAb is formulated in PBS. It should be stored at 4°C for short-term use and at -20°C for long-term storage. Aliquot the antibody upon receipt to avoid repeated freeze-thaw cycles, which can significantly reduce antibody performance.

Which secondary antibody should I use with ADAM9 Mouse mAb?

Because ADAM9 Mouse mAb is a mouse-derived monoclonal antibody, you should use an anti-mouse secondary antibody conjugated to the appropriate reporter (e.g., HRP for Western blot or DAB-based IHC, or fluorophore for immunofluorescence). Ensure the secondary antibody is raised in a species other than mouse to avoid cross-reactivity.

What is the expected molecular weight of ADAM9 detected by this antibody?

ADAM9 has a reported molecular weight of approximately 90–110 kDa depending on post-translational modifications and processing; however, the precise expected band size should be confirmed from the official product datasheet, as glycosylation and shedding events can alter apparent molecular weight on SDS-PAGE.

Can ADAM9 Mouse mAb be used for immunoprecipitation (IP)?

Suitability for immunoprecipitation has not been explicitly confirmed in the source material provided. Please refer to the current product datasheet or contact Aladdin Scientific technical support to confirm whether Ab351140 has been validated for IP applications.

References

1. Mochizuki S, Okada Y. ADAMs in cancer cell proliferation and progression. Cancer Sci. 2007;98(5):621-628.
2. Edwards DR, Handsley MM, Pennington CJ. The ADAM metalloproteinases. Mol Aspects Med. 2008;29(5):258-289.

Products are supplied for research and development use only. Not for use in humans, animals, diagnosis, or therapy.