Mouse IgG_2a Isotype Control [Clone C1.18.4] — Purified in vivo PLATINUM™ Functional Grade

Clone C1.18.4 is widely used in in vivo mouse studies as a mouse IgG2a isotype control antibody. Its primary function is to serve as a negative control to estimate non-specific binding of primary antigen-specific antibodies, ensuring that observed effects are due to specific antibody-antigen interactions and not background or off-target effects.

Key usage details and context:

• Isotype Control Purpose: C1.18.4 is structurally similar to experimental antibodies (same class and subclass: mouse IgG2a, kappa) but lacks specificity for mouse antigens. This allows researchers to distinguish true biological effects of their test antibody from non-specific immune responses triggered by antibody presence itself.
• In Vivo Applications: It is specifically validated for use in live mice to control for non-specific immune activation in animal models. Applications include control for depletion studies (e.g., Gr-1+ myeloid cell depletion), as well as a negative control in flow cytometry and other immunodetection techniques using live or fixed mouse tissues.
• Specificity: The C1.18.4 clone was originally generated against keyhole limpet hemocyanin (KLH), which is not present in mouse tissues, ensuring minimal or no binding to endogenous mouse proteins.
• Low Endotoxin Grades: In vivo grade C1.18.4 isotype control preparations are produced with very low endotoxin levels (<1 EU/mg protein), as higher endotoxin contamination could otherwise activate innate immune responses and confound in vivo results.
• Negative Control in Assays: The antibody is routinely used in ELISA, Western blot, immunoprecipitation, immunohistochemistry, and especially in in vivo animal experiments as a control for mouse IgG2a antibodies.

In summary, in vivo mouse studies use clone C1.18.4 primarily as an isotype-matched negative control to accurately interpret the biological specificity and functional effects of experimental murine IgG2a antibodies in live animals.

C1.18.4 is most commonly used as a mouse IgG2a isotype control antibody, particularly in experiments requiring non-specific controls alongside antigen-specific IgG2a antibodies. The antibodies or proteins frequently used together with C1.18.4 depend on the experimental design and target of interest, but the following are commonly paired in published literature:

• Antigen-specific mouse IgG2a antibodies: C1.18.4 provides a control for the non-specific effects of IgG2a isotype antibodies, so it is routinely used alongside other mouse IgG2a antibodies designed to target specific antigens (e.g., anti-CD3, anti-PD-1, anti-ICAM-1).

• Other Isotype Controls: Common isotype controls used in parallel include mouse IgG1 (e.g., MOPC-21), mouse IgG2b (e.g., MPC-11), or mouse IgG3 (e.g., MG3-35), depending on the isotype of the test antibody.

• Antibodies for functional and phenotypic analysis: In flow cytometry, immunohistochemistry, or ELISA, C1.18.4 is used alongside panel antibodies that recognize cell surface or intracellular markers; examples include anti-CD4, anti-CD8, anti-CD19, and anti-FoxP3.

• Secondary antibodies: When C1.18.4 is used in indirect detection systems, it is commonly paired with anti-mouse IgG2a secondary antibodies conjugated to fluorescent dyes or enzymes for visualization.

• Experimental proteins or antigens: Because C1.18.4 is an anti-keyhole limpet hemocyanin (KLH) antibody, it is sometimes used with KLH or in control experiments where the antigen is absent, to show specificity of binding.

Below is a simple comparison table for context:

In summary, C1.18.4 is almost always used in conjunction with antigen-specific mouse IgG2a antibodies and often with other mouse isotype controls, cell marker antibodies, and appropriate secondary reagents, depending on the specific immunological assay.

Clone C1.18.4 is widely cited in scientific literature as a mouse IgG2a isotype control antibody, primarily used for experimental controls in immunology studies. Key findings from its citations include:

• Role as an Isotype Control:
  C1.18.4 is specifically employed as a negative control for mouse IgG2a antibodies in techniques such as flow cytometry, cell depletion assays, and in vivo experiments. It does not bind to any known mouse antigens, which ensures that observed experimental effects are due to the activity of the primary antibody, not nonspecific binding or background signal.

• Fc Muted Variant for Improved Specificity:
  Some forms of C1.18.4, such as the LALA-PG "Fc Muted" format, contain mutations in the Fc region to minimize or eliminate Fc receptor and C1q binding. This reduces antibody-dependent cellular cytotoxicity (ADCC) and phagocytosis, further ensuring it serves only as a control for nonspecific IgG2a background and not for effector functions.

• Applications Documented in Major Studies:
  C1.18.4 has been cited in studies investigating immune regulation, inflammatory disease, and cancer, such as:

  • Wright et al., 2000 & 2001: Used as an isotype control while studying proinflammatory cytokine signaling and the role of immune complexes in arthritis.

  • Nishikawa et al., 2000: Served as an isotype control in dermatology research to compare with active primary antibodies and validate specificity.

  • Ikeda et al., 2003: Utilized as a negative control for in vivo depletion studies in immune cell populations.

• Quality Control and Cross-Reactivity Testing:
  The clone has been validated for minimal cross-reactivity against a range of cells and tissues across mouse, rat, and human species, supporting its reliability as a universal isotype control.

• Common Use in Tumor Immunology:
  While C1.18.4 primarily acts as a background control, some publications on tumor immunity (e.g., studies of myeloid cell depletion or immune infiltrate measurement) reference its use to demonstrate the specific effects of antibody therapy independent of any non-specific effects from the antibody isotype alone.

In summary, citations of C1.18.4 consistently document its use as a rigorously tested negative control antibody in immunological experiments, ensuring experimental specificity and reliability in research on cell populations, cytokine signaling, and immune-mediated disease models.

The dosing regimens of clone C1.18.4 in mouse models are not standardized, as this antibody is used as an isotype control, and dosing typically matches that of the test antibody being studied in each experimental protocol. This means the dose, schedule, and route of administration for C1.18.4 vary based on the study design and the antibodies with which it is paired, rather than being tailored to particular mouse models or disease applications.

Key points regarding dosing regimens:

• Researchers are advised to use C1.18.4 at the same concentration and dosing schedule as the experimental (test) antibody for which it serves as an isotype control.
• No universal or recommended dose exists for C1.18.4 itself; it is adjusted based on the specifics of each experiment, which may differ between mouse strains, disease models, and primary antibody application.
• Dosing information published for other mouse IgG2a antibodies (e.g., anti-VEGF or anti-PD-L1 antibodies used at 5 mg/kg, 2×/week, intraperitoneally in preclinical studies) may serve as general guidance, but these do not establish a fixed regimen for C1.18.4 itself. Instead, C1.18.4 would be used at these same doses if matched to such primary antibodies.
• The most relevant variable is matching the dosing to that of the experimental antibody, to ensure that any biological effects observed are due to antigen specificity rather than the IgG2a backbone or dosing regimen.

Application:

• C1.18.4 is primarily used in immunological research, especially for flow cytometry and in vivo studies, to control for non-specific binding and immune activation resulting from the IgG2a isotype.
• It is rarely, if ever, tested for efficacy or biological activity on its own in mouse disease models, as it does not target any mouse antigen.

In summary, the dosing regimen for C1.18.4 is determined by the primary (test) antibody's protocol in each experimental mouse model, rather than any inherent characteristic of clone C1.18.4 itself. There is no standardized or model-specific dosing apart from this principle.