SECTION 1 — SPECIMEN COLLECTION, LAB HANDLING & GENERAL LAB FACTS

Role of the Clinical Laboratory

• Diagnosis begins with proper specimen collection.

• Lab performs microscopy, culture, molecular tests (PCR), antigen tests.

• Understanding lab workflow helps interpret preliminary vs. final results.

Requirements of a Good Specimen

• Good quality

• Right timing

• Proper transport

• Adequate volume

• Correct container

• Label: patient name, date, sample source

• Requisition form must match label

• Transport immediately; store according to guidelines if delayed.

Standard Precautions (CDC)

Assume all patients may carry transmissible infection.

Applies to:

• Blood

• Body fluids (except sweat)

• Secretions & excretions

• Mucous membranes

• Non-intact skin

Precautions:

• Avoid sharps injuries

• Hand hygiene

• PPE: gloves, masks, gowns, eye protection

• Specimens handled in biosafety cabinets

• Cultures treated as potential pathogens

Laboratory Safety

• No eating/drinking/smoking

• Wash hands before/after lab

• Disinfect benchtop

• Keep work area clean

• No contaminated tools on bench

• Tie back hair

• Stay calm and controlled

• Do not remove cultures

• If spill: cover with towel + disinfectant, report

• Microbes do not “fly”; controlled with technique

Normal Flora — General Principles

• Newborns are sterile before birth; colonized during labor.

• Normal flora is usually harmless or beneficial.

• Only superficial tissues are colonized (skin, mouth, GI, respiratory, genital tract).

• Influenced by age, diet, medication, lifestyle, environment, climate, clothing.

• Infection occurs when organisms enter normally sterile sites.

• Trauma (cuts, burns, needles, surgery) introduces pathogens.

Principles of Diagnostic Microbiology

• Goals: culture, identify microbes, use biochemical/immunologic/molecular methods.

• Appropriate media needed for isolation.

• Correct biochemical tests needed for identification.

• Even with molecular tests, culture behavior matters.

Specimen Collection & Transport (HIGH-YIELD)

General Rules

• Use aseptic technique.

• Perform hand hygiene.

• Wear gloves.

• Collect at optimal time.

• Sample must be representative (e.g., deep pus, not surface).

• Use sterile container.

• Keep outside clean.

• Label properly.

Transport

• Use paper bag or sealed plastic bag.

• Deliver immediately.

• If delay: store according to lab instructions.

Respiratory Specimen Concepts

• Sputum, not saliva, is the correct specimen.

• Gram stain assesses:
  

  • Sample quality

  • Presence of neutrophils

  • Predominant organism type

Indigenous Flora — Importance

• Predict infections after injury.

• Identify source of infection.

• Understand overgrowth.

• Appreciate immune stimulation.

Antimicrobial Susceptibility Testing (AST)

1. Prepare suspension to 0.5 McFarland

2. Swab Mueller-Hinton agar

3. Apply antibiotic disks

4. Incubate 18–24 hours

5. Measure zone diameters

6. Interpret as S/I/R

Blood Culture Collection

• Clean skin with 70% alcohol + chlorhexidine

• Volume: adults 10–20 mL; children 1–5 mL

• Number: at least 2 bottles, up to 4

• Timing:
  

  • Intermittent bacteremia → collect during chills/fever

  • Continuous → timing not critical

UTI Specimen Handling - sterility

• Urine in bladder/kidneys is sterile.

• Becomes contaminated at distal urethra during voiding.

UTI Specimen Handling - clean-catch technique

1. Wash hands

2. Open sterile container

3. Clean genitals (front → back)

4. Begin voiding

5. Collect midstream

6. Label properly

UTI Specimen Handling - avoiding errors

• Refrigerate if delayed

• Use boric acid tubes

• Minimize contamination

UTI specimen handling - chemical indicators

• Nitrite → Gram–negative rods

• Leukocyte esterase → WBCs

UTI specimen handling - quantitative culture

• Use calibrated loop

• Multiply colonies ×100

• 100,000 CFU/mL = significant

Virology

• Viruses are obligate intracellular parasites.

• Contain DNA or RNA, not both.

• Identified via:
  

  • Cell culture (CPE)

  • Direct antigen testing

  • PCR

  • Serology (IgM = acute; IgG = past)

Prions

• Infectious proteins only (no DNA/RNA).

• Cause slow, fatal neurodegenerative diseases.

• Extremely resistant to heat, radiation, proteases.

• Require strong disinfectants (NaOH, bleach).