Unit 2: Core biochemistry panel estimation
Clinical Biochemistry-1 notes · PTU syllabus (BMLS403-18)
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Unit summary
A core panel of tests assesses diabetes, kidney and liver function, lipids and electrolytes. This unit covers the principles and reference ranges of glucose, proteins, urea, uric acid, creatinine, bilirubin and lipids, and the estimation of sodium, potassium, chloride, iodine, calcium and phosphorus.
After this unit you can
- Explain the principles of glucose and protein estimation
- Explain urea, creatinine, uric acid and bilirubin assays
- Explain lipid estimation
- Estimate electrolytes, calcium, phosphorus and iodine and correct errors
PTU syllabus topics
- Principles of assay procedures and normal/reference ranges for glucose
- proteins
- urea
- uric acid
- creatinine
- bilirubin and lipids
- principles
- procedures and error correction for sodium
- potassium
- chloride
- iodine
- calcium and phosphorus estimation
Fasting glucose
70–100 mg/dL
Diabetes
Urea
15–40 mg/dL
Kidney failure, dehydration
Creatinine
0.6–1.2 mg/dL
Kidney failure
Uric acid
3.5–7.2 mg/dL
Gout
Total bilirubin
0.3–1.2 mg/dL
Jaundice
Sodium / potassium
135–145 / 3.5–5.0 mmol/L
Electrolyte imbalance
Topic 1
Glucose
Glucose oxidase–peroxidase (GOD-POD)
Glucose → gluconic acid + H₂O₂; peroxidase with 4-aminoantipyrine and phenol gives a pink dye (505 nm)
Most common; specific
Hexokinase
Glucose-6-phosphate formed; NADPH measured at 340 nm
Reference method
Folin–Wu and o-toluidine
Older reduction or condensation methods
Less specific
- Reference ranges: fasting 70–100 mg/dL; 2-hour post-prandial below 140 mg/dL; HbA1c below 5.7%. Collect in fluoride tubes; glycolysis lowers glucose about 5–7% per hour at room temperature in plain tubes.
Topic 2
Proteins
- Total protein — biuret method: peptide bonds react with cupric ions in alkali to give a violet colour (540 nm). Albumin — bromocresol green (BCG): dye binding at pH 4.2 (630 nm). Globulin = total protein − albumin.
- Reference: total protein 6.0–8.3 g/dL; albumin 3.5–5.2 g/dL; A/G ratio 1.2–2.2. Low albumin: liver disease, nephrotic syndrome, malnutrition; high globulin: myeloma, chronic infection.
Topic 3
Urea, creatinine and uric acid
Urea
Urease–GLDH (kinetic, 340 nm) or diacetyl monoxime (DAM)
15–40 mg/dL
Creatinine
Jaffe's (alkaline picrate, orange) or enzymatic
0.6–1.2 mg/dL (men); 0.5–1.1 (women)
Uric acid
Uricase–peroxidase
Men 3.5–7.2; women 2.6–6.0 mg/dL
- Jaffe's interferences: ketones, glucose and cephalosporins raise results; kinetic and enzymatic methods reduce error. eGFR is calculated from creatinine, age and sex (CKD-EPI). Raised urea: kidney failure, dehydration, GI bleeding; raised uric acid: gout, leukaemia treatment.
Topic 4
Bilirubin
- Van den Bergh (diazo) reaction: bilirubin + diazotised sulphanilic acid → azobilirubin (pink–purple). Direct (conjugated) reacts without accelerator; total reacts with an accelerator (caffeine or methanol); indirect = total − direct.
- Reference: total 0.2–1.2 mg/dL; direct up to 0.3 mg/dL. Protect samples from light (bilirubin breaks down).
Topic 5
Lipids
Total cholesterol
Cholesterol esterase–oxidase–peroxidase (CHOD-PAP)
Below 200 mg/dL
Triglycerides
Lipase–glycerol kinase–GPO–peroxidase (GPO-PAP)
Below 150 mg/dL (fasting)
HDL cholesterol
Precipitate other lipoproteins (phosphotungstate) or direct homogeneous assay
Above 40 (men), 50 (women)
LDL cholesterol
Friedewald calculation or direct assay
Below 100 mg/dL optimal
Topic 6
Sodium, potassium and chloride
Sodium
Ion-selective electrode (ISE); flame photometry
135–145 mmol/L
Potassium
ISE; flame photometry
3.5–5.0 mmol/L
Chloride
ISE; mercuric thiocyanate colorimetry; coulometric titration
98–107 mmol/L
- Haemolysis
- Falsely raises potassium — reject and recollect
- Delayed separation or refrigeration of whole blood
- K⁺ leaks from cells — separate within 1–2 hours
- EDTA contamination
- Falsely high K⁺, low Ca²⁺
- Drip-arm sampling
- Dilution by IV fluids
- Lipaemia and high proteins
- Pseudohyponatraemia with indirect ISE or flame photometry — use direct ISE
- Electrode maintenance
- Calibrate, clean, replace membranes
Topic 7
Calcium, phosphorus and iodine
Calcium
o-Cresolphthalein complexone (OCPC) or arsenazo III; ionised calcium by ISE
8.5–10.5 mg/dL (total)
Phosphorus
Phosphomolybdate (UV at 340 nm or reduced to molybdenum blue)
2.5–4.5 mg/dL (adults); higher in children
Iodine
Sandell–Kolthoff reaction (iodide catalyses reduction of ceric ions by arsenite) in urine
Urinary iodine 100–199 µg/L adequate (WHO)
Formula
Corrected Ca = measured Ca + 0.8 × (4.0 − albumin in g/dL)
Example
Calcium 8.0 mg/dL with albumin 2.5 g/dL: corrected = 8.0 + 0.8 × 1.5 = 9.2 mg/dL (normal).
- Errors: EDTA or oxalate tubes chelate calcium (use serum or heparin); haemolysis raises phosphorus; prolonged tourniquet raises calcium.
Key terms
- GOD-POD
- Enzymatic glucose oxidase–peroxidase method
- Biuret reaction
- Violet colour of peptide bonds with copper in alkali
- Jaffe's reaction
- Creatinine with alkaline picrate
- Van den Bergh reaction
- Diazo reaction for bilirubin
- Ion-selective electrode
- Electrode measuring a specific ion's activity
Quick revision
- GOD-POD, hexokinase; fluoride tubes; reference values.
- Biuret, BCG; A/G ratio.
- Urease, Jaffe's, uricase; eGFR.
- Diazo bilirubin direct and total; light protection.
- CHOD-PAP, GPO-PAP, HDL, LDL; ISE electrolytes and errors; calcium, phosphorus, iodine; corrected calcium.
Important exam questions
Practice questions written to the PTU exam pattern for this unit's syllabus: short answers (Section A style) and long answers (Sections B and C style).
Short-answer questions
- Q1.State the principle of the GOD-POD method.
- Q2.Why is fluoride used for glucose samples?
- Q3.Name the method for serum albumin.
- Q4.What interferes with Jaffe's creatinine method?
- Q5.Why is haemolysis a problem for potassium?
- Q6.Correct calcium of 7.6 mg/dL with albumin 3.0 g/dL.
Long-answer questions
- Q1.Describe the estimation of blood glucose.
- Q2.Describe kidney function tests: urea, creatinine and uric acid.
- Q3.Describe the estimation of serum bilirubin and proteins.
- Q4.Describe the estimation of electrolytes and sources of error.
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