---

Biochemical Tests - A Comprehensive Overview

---

1. Liver Function Tests (LFTs)

A. Hepatocyte Injury Markers (Chemical-Metabolic System)

B. Synthetic Function Markers

C. Biliary/Canalicular Markers

Six Classic LFT Patterns

• Henry's Clinical Diagnosis and Management by Laboratory Methods, pp. 139-141
• Harper's Illustrated Biochemistry, 32nd Ed

---

2. Renal Function Tests

A. Blood Urea Nitrogen (BUN)

• BUN is inversely proportional to GFR: BUN ∝ 1/GFR
• Reference range: 10-20 mg/dL
• Limitation: affected by hydration status, catabolic state, and diet

B. Serum Creatinine

• Reference range: ~0.5-1.0 mg/dL
• Filtered and excreted; serum creatinine rises as GFR falls
• Important: a 75% reduction in GFR may cause only a modest rise in creatinine (non-linear relationship - see figure below)

C. BUN/Creatinine Ratio (key diagnostic tool)

D. GFR & Creatinine Clearance

E. Urine Tests for Renal Function

• Henry's Clinical Diagnosis and Management by Laboratory Methods, pp. 137-138
• Barash's Clinical Anesthesia, 9e

---

3. Thyroid Function Tests (TFTs)

• TSH is the most sensitive and first-line test
• Free T4 and free T3 are preferred over total levels because they are unaffected by changes in binding proteins
• Harper's Illustrated Biochemistry, 32nd Ed

---

4. Adrenal Function Tests

---

5. Cardiac Biomarkers

• Harper's Illustrated Biochemistry, 32nd Ed; Tintinalli's Emergency Medicine

---

6. Ascitic Fluid Biochemistry (Body Fluid Tests)

• Tietz Textbook of Laboratory Medicine, 7th Ed

---

7. Quick Reference: Common Reference Ranges

---

---

Biochemical Tests for Microorganisms

---

The Core Principle

---

Gram-Positive Cocci - The Key Algorithm

---

The 14 Standard Biochemical Tests

1. Catalase Test

• Principle: Catalase enzyme converts H₂O₂ → H₂O + O₂ (visible bubbles)
• Method: Add a drop of 3% hydrogen peroxide to a colony on a glass slide
• Positive result: Rapid bubbling/effervescence
• Key use:
  • Differentiates Staphylococcus (catalase +) from Streptococcus (catalase -)
  • Also positive: Micrococcus, Campylobacter, Listeria, Bacillus
• Note: Do NOT use blood agar - blood can cause false positives

---

2. Coagulase Test

• Principle: Coagulase enzyme + plasma factor → converts fibrinogen to fibrin clot
• Two methods:

• Coagulase +: Staphylococcus aureus
• Coagulase -: S. epidermidis, other CoNS
• Note: S. lugdunensis is slide coagulase positive but tube coagulase negative

---

3. Oxidase Test

• Principle: Detects the cytochrome c oxidase enzyme component (part of electron transport chain)
• Method: Reagent changes from colorless → blue/purple when oxidized
• Positive result: Blue/purple pigment within seconds
• Key use: Initial classification of Gram-negative rods
  • All Enterobacterales (e.g., E. coli, Klebsiella, Salmonella, Shigella) are oxidase NEGATIVE
  • Oxidase POSITIVE organisms: Pseudomonas aeruginosa, Neisseria, Campylobacter, Vibrio
• This single test separates the entire family Enterobacterales from other Gram-negative rods

---

4. Urease Test

• Principle: Urease hydrolyzes urea → 2 NH₃ + CO₂; ammonia raises medium pH
• Result: Pink/red color change (alkaline shift)
• Organisms:
  • Rapid (within 4 hrs) urease positive: Proteus spp., H. pylori
  • Positive: Klebsiella, Helicobacter, Ureaplasma
  • Negative: E. coli, Salmonella
• Clinical use: CLO test (Campylobacter-Like Organism test) is the bedside rapid urease test for H. pylori on gastric biopsy

---

5. Indole Test

• Principle: Tests ability to split indole (a benzopyrrole) from tryptophan
• Detection: Addition of Kovacs' reagent (p-dimethylaminobenzaldehyde) → red ring at surface
• Positive organisms: E. coli, Proteus vulgaris
• Negative organisms: Klebsiella, Proteus mirabilis, Salmonella
• Can be done as a rapid spot test from isolated colonies in seconds

---

6. Methyl Red (MR) Test

• Principle: Detects stable acid end products from glucose fermentation via mixed acid pathway
• Method: Add methyl red indicator after glucose fermentation
• Positive result (red): Strong acid production - E. coli, Shigella
• Negative result (yellow): Little acid - Klebsiella, Enterobacter

---

7. Voges-Proskauer (VP) Test

• Principle: Detects acetoin (acetylmethylcarbinol), an intermediate in the butylene glycol fermentation pathway
• Method: Add alpha-naphthol + KOH → red color if acetoin present
• Positive organisms: Klebsiella, Enterobacter (VP+, MR-)
• Negative: E. coli, Shigella (VP-, MR+)
• The MR and VP tests are typically opposite for the same organism

Together with Indole and Citrate, MR and VP form the classic IMViC battery used to differentiate members of the Enterobacteriaceae family.

---

8. Citrate Utilization Test

• Principle: Tests ability to use citrate as the sole carbon source
• Medium: Simmons Citrate Agar (with bromothymol blue pH indicator)
• Positive result: Growth + blue color change (alkaline)
• Positive organisms: Klebsiella pneumoniae, Salmonella, Enterobacter, Citrobacter
• Negative organisms: E. coli, Shigella

---

9. Hydrogen Sulfide (H₂S) Production

• Principle: Some bacteria produce H₂S from sulfur-containing amino acids
• Detection: H₂S reacts with iron salts → black precipitate (ferrous sulfide)
• Positive organisms: Salmonella, Proteus, Citrobacter
• Negative: E. coli, Klebsiella
• Used in TSI (Triple Sugar Iron) agar along with fermentation reactions

---

10. Carbohydrate Fermentation Tests

• Principle: Acidic metabolic products from carbohydrate breakdown change pH indicators
• Sugars tested: glucose, lactose, sucrose, mannitol, maltose
• Key clinical example: Lactose fermentation differentiates coliforms from non-coliforms
  • Lactose fermenters (pink colonies on MacConkey): E. coli, Klebsiella
  • Non-fermenters (colorless on MacConkey): Salmonella, Shigella, Pseudomonas
• Gas chromatographic detection of short-chain fatty acids from glucose fermentation is useful for anaerobic bacteria classification

---

11. Nitrate Reduction Test

• Principle: Detects bacterial reduction of nitrate → nitrite (or N₂ gas)
• Positive organisms: Most Enterobacteriaceae, Pseudomonas
• Clinical use: Nitrite dipstick in urinalysis detects Gram-negative UTI pathogens

---

12. ONPG Test (O-Nitrophenyl-β-D-Galactoside)

• Principle: Tests for presence of β-galactosidase enzyme
• Identifies organisms that have β-galactosidase but lack lactose permease - so they appear lactose-negative on standard media but are ONPG positive
• Example: Late lactose fermenters like Shigella sonnei

---

13. Decarboxylase / Deaminase Tests (Lysine, Ornithine, Arginine)

• Principle: Amino acid decarboxylation produces amines → alkaline shift detected by pH indicator
• Key example: Lysine decarboxylase positive = Klebsiella, E. coli
• Used primarily with Gram-negative rods

---

14. PYR Test (Pyrrolidonyl Arylamidase)

• Principle: Rapid colorimetric test; positive = pink to red color change
• Key organisms: Identifies Group A Streptococci (S. pyogenes), Enterococci, and S. lugdunensis
• Fast, can give presumptive ID from colonies within minutes

---

IMViC Summary Table (Enterobacteriaceae Differentiation)

---

Oxidase-Based Gram-Negative Classification

---

Modern Automated Methods

---

Quick Organism-Test Summary

---

---

How to Get Cruickshank's Medical Microbiology Legally

1. Library Access

• Check your university or medical college library - most institutions hold physical or digital copies
• Ask your librarian for interlibrary loan if not available locally

2. Purchase

• Search on Amazon, AbeBooks, or local medical bookstores
• Second-hand copies are often available at low cost on AbeBooks

3. Online Legal Platforms

• Google Books - often has partial previews
• Open Library (Internet Archive) - older editions may be available for free borrowing
• WorldCat - find the nearest library holding a copy

4. Institutional Access

• If you are a student, check if your institution subscribes to ClinicalKey, AccessMedicine, or Elsevier platforms

---

Note: The full title is "Medical Microbiology" by Cruickshank, Duguid, Marmion & Swain. Older editions (e.g., 12th) may be available through Internet Archive for free borrowing as they are out of print.