Anti-Mouse MHC Class II (I-A) [Clone Y-3P] – Purified in vivo PLATINUM™ Functional Grade

Common In Vivo Applications of Clone Y-3P in Mice

Blockade of MHC Class II Function

Clone Y-3P is a monoclonal antibody targeting mouse MHC Class II (I-A) molecules, and its primary use in vivo is to block MHC Class II-mediated immune responses. By binding to I-A molecules on antigen-presenting cells, Y-3P prevents the interaction of MHC Class II with the T cell receptor (TCR), thereby inhibiting T cell activation and downstream immune responses. This blockade is a powerful tool for studying the roles of MHC Class II in various immunological processes.

Experimental Autoimmune Encephalomyelitis (EAE) Model

Y-3P has been specifically used in the experimental autoimmune encephalomyelitis (EAE) model, a common mouse model for multiple sclerosis. In this context, administration of Y-3P neutralizes I-A molecules, which is critical for the induction and progression of EAE, thereby allowing researchers to investigate the contribution of MHC Class II-restricted T cell responses to autoimmunity.

General Immunological Blockade Studies

Beyond EAE, Y-3P is employed in a wide range of in vivo studies aimed at understanding the function of MHC Class II in immune regulation, tolerance, and pathogen clearance. These applications include experiments where researchers wish to transiently suppress MHC Class II-dependent antigen presentation to elucidate mechanisms of immune activation or tolerance.

Technical Details

• Target Specificity: Y-3P reacts with multiple mouse MHC Class II haplotypes (I-Ab, I-Af, I-Ap, I-Aq, I-Ar, I-As, I-Au, I-Av) and weakly with I-Ak.
• Isotype: Mouse IgG2a, κ.
• Administration: Typically injected intraperitoneally or intravenously for in vivo blockade studies.
• Formulation: Usually provided in PBS without stabilizers or preservatives, with low endotoxin levels for in vivo use.
• Controls: Isotype-matched control antibodies are recommended to distinguish specific from non-specific effects.

Summary Table

Key Points

• Y-3P is a go-to tool for in vivo blockade of MHC Class II function in mice, enabling researchers to dissect the role of CD4+ T cell responses in health and disease.
• Its use in the EAE model highlights its utility in autoimmune research, where MHC Class II presentation is central to disease pathogenesis.
• The antibody is well-characterized, with established protocols for in vivo use and available from multiple commercial sources.

In summary, clone Y-3P is predominantly used in vivo to block MHC Class II function, thereby modulating CD4+ T cell responses in models of autoimmunity, infection, and general immunology.

Y-3P is a monoclonal antibody that recognizes mouse MHC Class II (I-A) molecules and is commonly used in immunological research. Several other antibodies and proteins are frequently employed alongside Y-3P in various experimental contexts.

MHC Class II-Specific Antibodies

Y-3P is often used in conjunction with other MHC Class II-targeting antibodies. The Y-Ae antibody (anti-Eα:IAb) is particularly notable, as it recognizes the Eα52-68 peptide bound to IAb MHCII molecules and has been instrumental in understanding central tolerance and alloreactive antigen presentation. Another antibody, RM5-112, shares similar reactivity patterns with Y-3P, binding to multiple I-A haplotypes.

The AF6-120.1 antibody is another IAb-specific reagent that has been used comparably to Y-3P in affinity studies. Additionally, researchers have developed novel peptide:MHCII-specific antibodies such as the W6 MAb (anti-2W:IAb), which was generated using advanced hybridoma methodology and demonstrates affinity comparable to established IAb antibodies including Y-3P.

Haplotype-Specific Control Antibodies

In blocking experiments and functional assays, Y-3P is frequently paired with haplotype-specific control antibodies. For instance, 10.2-16 and 3F12 antibodies have been used in blocking studies examining CD4 T cell responses to influenza peptides presented by I-As or I-Ak molecules. These experiments also employed negative control antibodies such as 14.4.4S (I-As negative control) and MKD6 (I-Ak negative control).

Flow Cytometry Markers

When used in flow cytometry applications, Y-3P is commonly combined with markers for antigen-presenting cell subsets. Studies frequently incorporate antibodies against CD11c, CD11b, and CD19 to identify specific dendritic cell populations. These combinations allow researchers to characterize MHC class II-expressing cells and assess antigen presentation in various experimental models.

Clone Y-3P is a monoclonal antibody against mouse MHC Class II (I-A) that has been extensively utilized in immunological research. The antibody has generated significant findings across multiple areas of immune function and regulation.

MHC Class II Blocking and TCR Stimulation

Clone Y-3P serves as a critical tool for in vivo blockade of T cell receptor (TCR) stimulation and MHC class II blocking, which are fundamental to understanding immune responses and antigen presentation. The antibody has been reported to inhibit I-A-restricted T cell responses, making it valuable for studying T cell activation mechanisms and immune regulation pathways.

Detection and Characterization of MHC Class II Expression

The Y-3P antibody has proven essential for detecting and characterizing MHC class II molecules in various experimental contexts. It has been used to determine MHC class II expression on B cells by staining splenocytes, and researchers have employed it to confirm bona fide MHCII expression in bone marrow-derived mast cells (BMMCs). The antibody successfully detects IA-specific molecules and has been used alongside other monoclonal antibodies like 10-2.16 and 11-5.2 to identify substantial amounts of class II MHC-peptide complexes on immature dendritic cells.

Biochemical Studies of MHC Class II Dynamics

Clone Y-3P has facilitated detailed biochemical analyses of MHC class II molecule behavior. In pulse-chase experiments, researchers have used Y-3P for immunoprecipitation of pulse-labeled and biotinylated MHC II molecules, followed by purification with streptavidin-agarose. These studies have enabled quantitative analysis of MHC II molecule turnover and stability using labeled proteins extracted with NP-40 buffer.

Functional Studies in Knockout Models

The antibody has been instrumental in characterizing mice lacking conventional MHC class II genes. In studies of MHCII knockout mice, Y-3P (anti-Ab antibody) was used alongside the broadly reactive anti-A/E reagent M5/114 to determine expression patterns on B cells. These experiments helped establish that MHCII knockout mice are viable and fertile with no obvious anatomical or behavioral defects.

Applications in Antibody Development

Clone Y-3P has supported advances in monoclonal antibody generation techniques. Research comparing novel methodologies for generating antibodies specific for peptide bound to MHCII has demonstrated significantly improved efficiency rates, with success rates ranging from 1:18 to 1:115 compared to traditional approaches of 1:250 to 1:1,000. This represents a 2.25 to 33.75-fold improvement in efficiency.

Dosing regimens for clone Y-3P antibodies in mouse models are highly variable and depend on several experimental parameters, including the mouse strain, targeted MHC-II haplotype, desired immunological effect (e.g., depletion vs. blockade), and route of administration.

Key considerations determining dosing strategies:

• Mouse strain and MHC-II haplotypes: Clone Y-3P reacts with multiple MHC class II haplotypes (I-Ab, I-Af, I-Ap, I-Aq, I-Ar, I-As, I-Au, I-Av, and weakly with I-Ak), necessitating adjustment of the dose to match the expression level and distribution of these molecules in the chosen mouse model.
• Experimental goal: The dose may differ if the aim is complete functional blockade of MHC-II-TCR interactions or mere modulation of immune responses. For inhibition studies, high single doses are sometimes employed, while for sustained immunomodulation, lower repeated doses may be favored.
• Administration route: Most protocols use intraperitoneal injection, but intravenous or subcutaneous routes may be used to achieve different pharmacokinetics.
• Typical regimens (as inferred from similar anti-MHC antibody protocols):
  • Doses often range between 100–250 μg per mouse, administered once or on a schedule (e.g., every 3-4 days) for several doses, to maintain blockade. Exact values for Y-3P should be confirmed from specific published experiments, as most guides provide ranges for similar clones.
  • Some studies utilize single higher doses for acute functional blockade, while others use repeated dosing for sustained effects.

Practical guidance:

• Pilot titration is recommended, as published optimal doses for Y-3P are model-dependent and should be calibrated according to the activity observed in the specific strain and experiment.
• Always refer to peer-reviewed literature or product-specific guidelines for the most appropriate dose and schedule—there is no universal standard.

In summary, dosing regimens for clone Y-3P in mouse models must be customized according to mouse genetics, experimental design, and intended immunological outcome. Standard doses are typically in the 100–250 μg/mouse range, but optimal scheduling and administration routes require experimental validation.