Anti-Mouse ICOSL (CD275) (Clone HK5.3) – Purified in vivo PLATINUM™ Functional Grade

Clone HK5.3 is a rat monoclonal antibody that specifically binds to mouse ICOSL (Inducible T-cell Co-Stimulator Ligand, also known as CD275, B7RP-1, or B7-H2) and is widely used in in vivo mouse studies to block the interaction between ICOSL and its receptor ICOS.

Key uses in in vivo mouse studies:

• Blocking ICOS–ICOSL Pathway: HK5.3 is administered to mice to prevent ICOSL from binding to ICOS on T cells. This blockade alters T-cell co-stimulation and immune responses.
• Immune Modulation: Treatment with HK5.3 leads to reduced regulatory T cell (Treg) numbers in vivo, impacting T cell and B cell function, as well as overall immune homeostasis.
• Disease Models: HK5.3 is used in models of autoimmune and inflammatory disease (such as the collagen-induced arthritis model) to suppress immune responses—specifically inhibiting both Th1 and Th2 responses and ameliorating disease severity.
• Experimental Techniques: The antibody is injected intraperitoneally or intravenously in mice to achieve systemic ICOSL neutralization, and isotypes or chimeric variants are available for specific experimental needs.

Summary table of HK5.3 use in mouse models:

Supporting Details:

• HK5.3 is a rat IgG2a antibody, and mouse IgG2a chimeric versions are also available for experiments requiring reduced immunogenicity.
• Its use is well characterized in both basic research and preclinical therapy studies related to immune regulation and checkpoints.

References to antibody suppliers and characterization can be found in Bio X Cell, Fisher Scientific, and primary literature.

The correct storage temperature for the sterile packaged HK5.3 clone (Anti-Mouse ICOSL/CD275 antibody) is 2–8°C for short-term use (up to four weeks); for long-term storage, aliquot and store at –80°C.

• For up to four weeks: 2–8°C (refrigerator), undiluted and avoiding repeated freeze-thaw cycles.
• For long-term storage: –80°C, in aliquots to prevent loss of activity.

Do not freeze at moderate temperatures (e.g., –20°C), as ultra-low temperatures (–80°C) provide the best stability for extended periods. Always consult the product’s Certificate of Analysis for incremental details if available.

The HK5.3 antibody is a monoclonal antibody targeting mouse ICOSL (CD275, also known as B7RP-1 or B7-H2), which is an immune checkpoint protein expressed on antigen-presenting cells. In the literature, researchers frequently use HK5.3 in combination with other antibodies or proteins to study T cell co-stimulation, immune regulation, and cell profiling in various immunological contexts.

Commonly used antibodies or proteins with HK5.3 include:

• Anti-ICOS (CD278) antibodies: Since HK5.3 blocks ICOSL, studies often pair it with anti-ICOS antibodies to evaluate both the ligand and receptor sides of the ICOS/ICOSL interaction, which is crucial for T and B cell co-stimulation.
• Anti-CD3 and anti-CD28 antibodies: These are commonly used for general T cell activation and costimulation studies in conjunction with modulation of the ICOS/ICOSL pathway.
• Anti-CD4 and anti-CD8 antibodies: These markers allow for the identification and profiling of T helper and cytotoxic T cell subsets when analyzing the effects of ICOSL blockade.
• Anti-Foxp3 or anti-CD25 antibodies: These are often used to identify or track regulatory T cells (Tregs), as HK5.3 treatment has been reported to lead to Treg loss, making co-staining for these markers relevant for mechanistic studies.
• Other B7 family antibodies (e.g., anti-CD80, anti-CD86): Researchers often investigate multiple immune checkpoints in parallel, using antibodies against other co-stimulatory molecules expressed on antigen-presenting cells.
• Isotype control antibodies: To ensure the specificity of HK5.3-related findings, matching isotype controls are routinely used in all antibody-based experiments.

Supporting Details:

• Experimental protocols employing HK5.3 often include panels of surface and intracellular markers to fully characterize the effects on immune cell subsets after ICOSL blockade.
• In vivo experiments typically combine HK5.3 with antibodies for cell depletion or tracing, immune checkpoint analysis, and cytokine detection to dissect complex immune responses.

Summary Table: Frequently Combined Antibodies/Proteins with HK5.3

In summary, HK5.3 is most commonly used with antibodies that (1) define immune cell lineages, (2) evaluate the impact on immune checkpoint pathways, and (3) control for non-specific effects.

Clone HK5.3 is a rat anti-mouse ICOSL monoclonal antibody that has been used in research to study immune responses and inflammatory conditions. Here are the key findings from its citations:

1. Immune Response Inhibition: HK5.3 is known to inhibit both Th1 and Th2 immune responses. This is significant because it can modulate the immune system's response in conditions where these pathways are involved.

2. Arthritis Amelioration: In the context of inflammatory arthritis, HK5.3 has been shown to ameliorate symptoms. This suggests potential therapeutic applications in managing arthritis or similar inflammatory conditions.

3. Comparison with Novel Therapies: In studies comparing HK5.3 with new anti-ICOSL therapies, such as VNAR-Fc domains, HK5.3 served as a control. These novel therapies showed comparable or improved efficacy in reducing inflammatory markers and tissue damage.

4. Treatment of Immune Responses: In experiments involving blockade of ICOS-L (CD275), HK5.3 was used to treat mice before and after immunization. This blockade impacts T cell and B cell responses, affecting germinal center formation and antibody production.

Overall, HK5.3 plays a crucial role in research related to immune modulation and the study of ICOSL's role in immune responses.