Related Experiment Video
Updated: Jan 18, 2026
Identification and Analysis of Mouse Erythroid Progenitors using the CD71/TER119 Flow-cytometric Assay
Published on: August 5, 2011
Epitope analysis of an anti-mouse CCR1 monoclonal antibody S15040E using flow cytometry
Ayaka Okada1, Hiroyuki Suzuki1, Takao Arimori2
1Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, 2-1 Seiryo-machi, Aoba-ku, Sendai, Miyagi, 980-8575, Japan.
Abstract:
The C-C motif chemokine receptor 1 (CCR1) is widely expressed in various immune cells and plays crucial roles in the maturation and migration of immune cells. CCR1 has been considered an attractive drug target for treating autoimmune diseases and tumors. An anti-mouse CCR1 (mCCR1) monoclonal antibody (clone S15040E) has been used in various in vivo studies to identify mCCR1-positive cells by flow cytometry. However, the binding epitope has not been determined. This study investigated the binding epitope of S15040E using flow cytometry. The mCCR1 extracellular domain-substituted mutant analysis showed that S15040E recognizes the extracellular loop 2 (ECL2, aa 172-197) of mCCR1. Next, alanine (or glycine) scanning was conducted in the ECL2 region. The results revealed that Trp176, Phe178, and Arg181 are essential amino acids for the recognition by S15040E. These results showed the involvement of the ECL2 of mCCR1 in the recognition by S15040E.
Insights
This study identified the specific binding site of an anti-mouse CCR1 antibody (S15040E). The antibody targets the extracellular loop 2 (ECL2) of mouse CCR1, crucial for immune cell function.
Area of Science:
- Immunology
- Molecular Biology
- Drug Discovery
Background:
- C-C motif chemokine receptor 1 (CCR1) is vital for immune cell function and a target for autoimmune diseases and tumors.
- An anti-mouse CCR1 (mCCR1) monoclonal antibody (clone S15040E) is used in vivo but its binding epitope is unknown.
Purpose of the Study:
- To determine the binding epitope of the anti-mCCR1 monoclonal antibody S15040E.
- To characterize the specific amino acid residues involved in antibody recognition.
Main Methods:
- Flow cytometry was used to analyze antibody binding.
- Extracellular domain-substituted mutant analysis of mCCR1 was performed.
- Alanine (or glycine) scanning of the mCCR1 ECL2 region was conducted.
Main Results:
- The S15040E antibody recognizes the extracellular loop 2 (ECL2, amino acids 172-197) of mCCR1.
- Specific amino acids Trp176, Phe178, and Arg181 within ECL2 are essential for S15040E binding.
- These findings confirm the critical role of the mCCR1 ECL2 in antibody recognition.
Conclusions:
- The binding epitope of the anti-mCCR1 antibody S15040E has been precisely mapped to the ECL2 region.
- Key amino acid residues (Trp176, Phe178, Arg181) within ECL2 are critical for antibody recognition.
- This detailed epitope information is valuable for understanding mCCR1 function and developing targeted therapies.
