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#antigen

3 public questions tagged with this topic.

Immunological Applications

Rituximab targets which antigen?

MS4A1 gene on chromosome 11q12.2 encodes CD20 differentiation antigen, 33 to 37 kDa tetra-spanning membrane phosphoprotein with both N and C termini intracellular, four transmembrane helices forming tetrameric oligomer functioning as store-operated calcium channel component modulating B cell receptor induced calcium flux via interaction with calcium release-activated calcium modulator ORAI1. Expression tightly regulated during ontogeny: absent on hematopoietic stem cells and early pro-B, appears at late pro-B stage after productive immunoglobulin heavy chain rearrangement, persists through mature naive, germinal center, memory B cells, but extinguished upon plasma cell differentiation when transcription factors BLIMP1 and XBP1 repress MS4A1 and induce secretory program. Lack on stem cells ensures regenerative capacity after depletion; absence on long-lived plasma cells preserves humoral memory maintaining protective antibody titers against prior vaccines. Rituximab binding extracellular loops between transmembrane domains 3-4 induces translocation to lipid rafts enriched in cholesterol and sphingolipid, enhancing Lyn mediated PLCγ2 activation, calcium mobilization, and Bax mediated mitochondrial apoptosis. Concurrent opsonization promotes phagocytosis by macrophages via FcγRI and complement. Reconstitution from pro-B precursors occurs within six to twelve months, infection risk transient compared to continuous pan-B ablation.

Ref: NCBI Gene MS4A1 CD20 tetra-span; Janeway Immunobiology CD20 calcium channel rituximab ADCC CDC.

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Basic Applied Biology

Rituximab targets antigen:

Rituximab represents first chimeric anti-cancer monoclonal antibody approved 1997 targeting CD20, a 33 to 37 kDa non-glycosylated tetra-span membrane phosphoprotein encoded by MS4A1 gene on chromosome 11q12 expressed on pre-B through mature B lymphocytes but absent on hematopoietic stem cells, pro-B cells and terminally differentiated plasma cells. Protein architecture comprises four transmembrane domains with short intracellular termini and two extracellular loops accessible for antibody binding. Physiologic function involves regulation of calcium flux through modulating B cell receptor signaling threshold, affecting activation and differentiation. Rituximab binding via Fab region to CD20 extracellular loop triggers multiple effector mechanisms: complement-dependent cytotoxicity via C1q recruitment and membrane attack complex formation, antibody-dependent cellular cytotoxicity mediated by Fc region engaging Fc-gamma-RIIIa on natural killer cells releasing perforin, direct apoptosis induction via cross-linking, lipid raft clustering and activation of caspase 3, and phagocytosis by macrophages. Specific lineage restriction allows B cell depletion in non-Hodgkin lymphoma, chronic lymphocytic leukemia, rheumatoid arthritis with subsequent regeneration from stem cells lacking CD20. Absence on other lineages minimizes off-target toxicity compared to CD4, CD8 or HER2 targets.

Ref: Maloney et al. Blood 1997 Rituximab ideal target; Reff et al. CD20 biology; PubMed review CD20 therapeutic target.

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Flow cytometry, FACS, ELISA

What happens to radioactive signal in RIA if antigen is high?

Radioimmunoassay principle involves fixed limited antibody, fixed trace amount of radiolabeled antigen, and variable unlabeled antigen from sample competing for binding. At equilibrium, higher concentration of unlabeled antigen occupies more antibody sites, displacing labeled antigen into free fraction that is washed away. Radioactivity measured in bound pellet therefore decreases as sample antigen increases. This inverse relationship allows construction of standard curve. No change or direct proportionality would violate competition kinetics, equal distribution would indicate no affinity. Bound counts fall with rising antigen levels.

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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