Anti-Rat CD28 (Clone JJ319) – Purified in vivo PLATINUM™ Functional Grade

Use of Clone JJ319 in In Vivo Mouse Studies

JJ319 is a monoclonal antibody that specifically recognizes rat CD28, a critical costimulatory receptor expressed on T lymphocytes and some NK cells. While its primary applications are well established in in vitro assays (e.g., flow cytometry, T cell stimulation, and immunoprecipitation), there is limited direct evidence of its use for in vivo administration in mice based on the available search results. Here’s a structured summary:

Key Properties

• Specificity: Anti-rat CD28 (clone JJ319).
• Host species: Mouse (mouse IgG1, ?).
• Species reactivity: Rat.
• Mechanism: JJ319 is a “conventional” anti-CD28 antibody, meaning it can costimulate T cell proliferation and cytokine production only when T cell receptors (TCR) are also engaged (e.g., by anti-TCR antibodies).
• In vivo context: Most sources describe JJ319’s use for in vitro stimulation or as a functional grade reagent for in vivo studies in rats. There is no explicit documentation in the provided results that JJ319 is routinely administered in vivo in mice—notably, the immunogen is A20J cells transfected with rat CD28, and product descriptions emphasize rat reactivity.

Comparison with Superagonistic Antibody JJ316

• JJ319 (conventional): Requires TCR engagement to deliver a costimulatory signal; does not directly activate T cells alone.
• JJ316 (superagonistic): Can directly activate T cells in vitro and in vivo without TCR engagement, making it a tool for potent T cell stimulation in both rats and mice.
• In summary: JJ316 is the antibody of choice for in vivo T cell activation in rodents (including mice), whereas JJ319 is primarily used for in vitro or rat in vivo studies where costimulation (not direct activation) is desired.

Practical Use in In Vivo Studies

• Rat studies: JJ319 is available in a functional grade formulation suitable for in vivo administration in rats.
• Mouse studies: No direct evidence from the provided sources supports the in vivo use of JJ319 in mice. Its cross-reactivity with mouse CD28 is not established in the product literature, and its primary described use is for rat CD28.
• In vitro use in mice: Possible (e.g., for stimulation of mouse T cells expressing rat CD28), but not the intended or common application.

Conclusion

Clone JJ319 is a tool for studying rat CD28, mainly used in vitro or in functional grade formulations for in vivo rat experiments. It is not standardly used for in vivo administration in mice; for mouse in vivo T cell studies, superagonistic anti-CD28 antibodies (such as clone JJ316 or hamster anti-mouse CD28 mAb 37.51) are typically employed instead. Always verify cross-reactivity and consult original publications or vendor data sheets before using anti-rodent antibodies in a non-native species.

For sterile packaged clone JJ319, a mouse anti-rat CD28 antibody, the correct storage temperature is 2–8°C (refrigeration) when the antibody is in solution (liquid form). The solution should be stored undiluted and protected from prolonged exposure to light. Aliquoting is recommended to avoid repeated freeze-thaw cycles, and the vial should be gently mixed before use. Do not freeze the antibody.

If you are referring to the antibody in a dry or lyophilized form, no specific storage temperature is provided for clone JJ319 in the available results, but for most antibodies in this state, storage at -20°C (freezer) is typical—however, always refer to the manufacturer’s datasheet for confirmation, as this was not explicitly stated in the search results.

Summary Table

Key Points

• Liquid antibody: 2–8°C, protected from light, do not freeze.
• Lyophilized antibody: Temperature not specified in results; consult manufacturer if in doubt.
• Avoid repeated freeze-thaw cycles for liquid forms to maintain antibody integrity.

In the literature, JJ319 is often used alongside other antibodies and proteins that target various components of the immune system, particularly those involved in T cell activation and signaling. Some of these include:

• Anti-mouse CD28 (clone 37.51): This antibody is used to study mouse CD28, acting similarly to JJ319 but with specificity for mouse CD28. It binds near the B7 binding site, similar to JJ319, but with different epitope recognition.

• Anti-human CD3 (clone UCHT1): This antibody is used to study human T cell activation and is often combined with CD28 antibodies for co-stimulation studies.

• Anti-human TCR? (clone 6B10.2) and anti-human ZAP-70 (clone 3.3.1): These antibodies are used to study T cell receptor signaling components.

• Phytohemagglutinin (PHA): Although not directly mentioned with JJ319, PHA is a common reagent used in T cell activation studies and could potentially be used in conjunction with JJ319 for immune response analysis.

JJ319 itself is a monoclonal antibody that recognizes rat CD28, a costimulatory receptor involved in T cell activation. It is used to study immune responses, particularly in rat models, and has been shown to induce profound immunosuppression in transplantation models.

Key Findings Related to Clone JJ319 in Scientific Literature

Epitope Recognition and Functional Differences

• JJ319 is a conventional monoclonal antibody (mAb) raised against rat CD28, distinguishing it from superagonists like JJ316, which activate T cells in a different manner.
• JJ319 binds to an epitope located between amino acids 66 and the transmembrane domain of rat CD28. Specifically, a critical residue for JJ319 binding is valine at position 98 in the extracellular region. This epitope is distinct from the region recognized by superagonists, which map to the C?D loop in the middle of the extracellular domain.
• The binding site for JJ319 is close to the B7 (CD80/CD86) binding site, suggesting that its mechanism of action could modulate natural ligand (B7) interactions, unlike superagonists that act independently of the B7-CD28 pathway.

Functional Effects

• Administration of JJ319 (0.1 mg) did not affect disease manifestation in experimental autoimmune encephalomyelitis (EAE) models, with clinical scores comparable to control-treated animals. This contrasts with the protective effects seen with the superagonist JJ316, highlighting the functional distinction between conventional and superagonistic anti-CD28 antibodies.
• There is no evidence that JJ319 induces robust T cell activation or expansion of regulatory T cells (Tregs), which is a hallmark of CD28 superagonist activity. This further emphasizes that JJ319 acts via a conventional, non-superagonistic mechanism.

Comparative Insights

• Binding of JJ319 is species-specific: It does not bind mouse CD28 unless specific amino acid changes (e.g., F98V) are introduced, illustrating the precise structural requirements for its recognition.
• JJ319 and the mouse CD28 mAb 37.51 recognize overlapping but distinct epitopes: JJ319 binding is lost when phenylalanine at position 98 is present (mouse), whereas 37.51 binding is lost when valine is present (rat), demonstrating reciprocal epitope requirements.

Summary Table: JJ319 vs. JJ316 (Superagonist)

Conclusion

Clone JJ319 is a conventional anti-rat CD28 monoclonal antibody that binds near the B7 interaction site and does not induce robust T cell activation or disease modulation in vivo. Its binding specificity and functional inactivity contrast sharply with CD28 superagonists, underscoring the importance of epitope location in determining the biological effects of anti-CD28 antibodies.