Immunology

Immune System Components

Innate Immunity (Non-specific)

The innate immune system provides the first line of defense and responds rapidly (minutes to hours) to pathogens without prior sensitization.

Adaptive Immunity (Antigen-specific)

The adaptive immune system develops over days and provides long-lasting protection through immunological memory.

Antibody Biology

Antibody Structure

Immunoglobulins (Ig) are Y-shaped proteins composed of two heavy chains and two light chains.

       ┌──────────────────────────────────────┐
       │           Variable Region            │
       │         (VH - Heavy Chain)           │
       │         (VL - Light Chain)           │
       └──────────────────┬───────────────────┘
                        ╱ ╲
       ┌────────────────┴───┴────────────────┐
       │            Hinge Region              │
       └──────────────────┬───────────────────┘
       ┌──────────────────┴───────────────────┐
       │         Constant Region (CH)        │
       └──────────────────────────────────────┘
              ┌──────────────┐
              │   Light      │
              │   Chain      │
              │   (CL)       │
              └──────────────┘

Antibody Classes (Isotypes)

Antibody Naming Conventions

• Polyclonal antibodies: Multiple epitopes, heterogeneous preparation
• Monoclonal antibodies: Single epitope, homogeneous, produced by hybridomas
• Nomenclature: Prefix + target + -mab (e.g., pembrolizumab, rituximab)
• Humanized antibodies: Murine variable regions grafted to human framework

T-Cell Immunology

T-Cell Receptor (TCR) Biology

The TCR is a heterodimer (αβ or γδ) that recognizes antigens presented on MHC molecules.

TCR Complex:
┌─────────────────────────────────────────┐
│         αβ Heterodimer (TCR)            │
├─────────────────────────────────────────┤
│         CD3 Complex (Signal Transduction)│
│         ζ-chain (homodimer)            │
├─────────────────────────────────────────┤
│         CD4 or CD8 Coreceptor            │
└─────────────────────────────────────────┘

Major Histocompatibility Complex (MHC)

T-Cell Differentiation States

Cytokines & Immune Signaling

Key Cytokines

Signaling Pathways

# Cytokine signaling conceptual model
class CytokineSignaling:
    """Simplified cytokine signaling pathway model"""
    
    JAK_STAT_PATHWAY = {
        "Type I (e.g., IL-6)": {
            "receptor": "gp130 family",
            "JAKs": "JAK1, JAK2, TYK2",
            "STATs": "STAT1, STAT3",
            "outcome": "Acute phase response, B cell differentiation"
        },
        "Type II (e.g., IFN-γ)": {
            "receptor": "IFNAR family", 
            "JAKs": "JAK1, JAK2",
            "STATs": "STAT1, STAT2",
            "outcome": "MHC upregulation, antiviral response"
        }
    }
    
    NFκB_PATHWAY = {
        "canonical": {
            "activators": ["TNF-α", "IL-1β", "LPS", "TLRs"],
            "IKK_complex": "IKKα, IKKβ, IKKγ",
            "inhibitor": "IκBα",
            "transcription_factor": "p65/p50"
        }
    }

Flow Cytometry Analysis

Sample Preparation

1. Cell count and viability: Aim for >90% viability, 1×10⁶ cells/mL
2. Staining: Surface markers (4°C, 30 min) → intracellular (fix/perm)
3. Controls:
   • Unstained control (autofluorescence)
   • Fluorescence minus one (FMO)
   • Isotype controls
   • Viability dye

Common Fluorophores

Gating Strategy Example

Raw Data
    ↓ (FS/SS gate) → Lymphocyte gate
    ↓ (Single cells) → Live/Dead exclusion
    ↓ (CD45+) → Leukocyte population
    ↓ (CD3+) → T cells
    ↓ (CD4/CD8) → Helper vs Cytotoxic
    ↓ (CD45RA/CCR7) → Naive vs Memory

Autoimmunity & Immunodeficiency

Autoimmune Disease Classification

Primary Immunodeficiencies

Vaccine Immunology

Vaccine Types

Vaccine Response Evaluation

# Serological response interpretation
class VaccineResponse:
    """Interpret vaccine-induced immune responses"""
    
    def calculate_seroconversion(self, pre_titer, post_titer, threshold=4):
        """4-fold rise = seroconversion"""
        if pre_titer < threshold:
            return post_titer >= threshold
        return post_titer / pre_titer >= 4
    
    def interpret_protective_titer(self, antibody_level, protective_level=10):
        """Assess if titer exceeds protective threshold"""
        return antibody_level >= protective_level
    
    # Memory B cell response (flow cytometry)
    def memory_bcell_response(self, antigen_specific_bcells, total_bcells):
        """Calculate percentage of antigen-specific memory B cells"""
        if total_bcells == 0:
            return 0
        return (antigen_specific_bcells / total_bcells) * 100

Immunological Techniques

ELISA (Enzyme-Linked Immunosorbent Assay)

Western Blot

• Purpose: Detect specific proteins in a mixture
• Detection: Primary antibody + enzyme-conjugated secondary antibody
• Substrate: Chemiluminescent (ECL), colorimetric, fluorescent

Common Errors to Avoid

1. Ignoring Fc receptor binding — use Fc block for phagocyte staining
2. Not accounting for dead cells — always include viability dye
3. Insufficient controls — FMO is essential for proper gating
4. Citing antibody clone without validation — verify specificity
5. Assuming isotype controls match primary — they should match fluorophore
6. Over-gating rare populations — report parent population percentages
7. Confusing correlation with causation — functional assays validate phenotypic findings