AL

Clinical Biochemistry – Quick Notes on Lab Tests & Sources of Error

From request to report

  • Clinical question → biochemical answer; request form includes clinical data; preanalytics → analytics → postanalytics; reporting including interpretation.
  • Purpose of lab tests: screening, diagnosis, monitoring, prognosis.
  • Major workflow idea: From request to report
    • Preanalytics account for the largest share of errors; analytics and postanalytics contribute less.
    • Quality depends on correct samples and proper handling at each step.

Phases of Laboratory Testing

  • Three phases: Preanalytical, Analytical, Postanalytical.
  • Quality focus: ensure reliable results to support correct medical decisions. "Garbage in, garbage out" if samples are faulty.
  • Error shares (approximate): preanalytical ~70–80%, analytical ~5–10%, postanalytical ~10–15%.

Pre-analytical: Patient-related factors

  • Diet and fasting: 12-hour fasting recommended (water allowed); lipemia affects photometric assays; reference ranges assume fasting state.
  • Alcohol: avoid in previous 24 h; caffeine and smoking in the morning can affect results.
  • Exercise and stress: extreme activity and short-term stress raise CK, cortisol, etc.; rest ~15 minutes recommended before draw.
  • Diurnal variation: cortisol highest in morning; iron highest at noon; TSH highest in evening; use morning samples for comparison.
  • Posture effects: standing vs seated changes in RBC, Hb, Hct, and other parameters.
  • Pregnancy: hemodilution (Hb ↓); ALP ↑; hypercoagulability.
  • Age and gender: reference ranges vary with age/gender; ALP higher in children; other parameters shift with development.
  • Medications and substances: morning meds; anticoagulants; acetaminophen → ↑GGT; vitamin C can affect some assays; smoking influences several markers.
  • Infections/illness: acute illness can skew iron, thyroid hormones, etc.; avoid during acute phases when interpreting certain tests.

Pre-analytical: Sample-related factors

  • Sample types: venous (most common), arterial (blood gas), capillary (infants/POCT).
  • Common preanalytical issues: infusion/dilution, parenteral nutrition–induced lipemia, cannula contamination with anticoagulants, contrast media effects; capillary sampling should avoid squeezing; warm the skin before collection.
  • Time sensitivity: certain samples require rapid handling (e.g., blood gases within 30 minutes).
  • Anticoagulants and container effects: wrong tube can alter results; proper blood-anticogulant ratio is essential.

Blood collection tubes and order of draw

  • Major tube types:
    • Light blue: citrate (coagulation tests).
    • Black: citrate (ESR).
    • EDTA (purple): hematology (CBC).
    • Heparin (green): chemistry.
    • Gray: glycolysis inhibitor.
  • Order of draw (typical): microbiology samples → citrate (coagulation) → ESR (citrate/black) → plain serum (gel/clot activator) → heparin → EDTA → gray.
  • Tubes may be pre-barcoded; ensure proper labeling, fill line visibility, and correct orientation.
  • Mixing: invert tubes gently (usually 5–10 times) to mix anticoagulant.

Sample handling and transport

  • Proper labeling and patient identification are crucial (name + date of birth or health ID).
  • Transport conditions and timing: prompt delivery to lab; some samples require immediate processing (e.g., CSF, ABG) or cooling on ice.
  • Avoid vigorous shaking; avoid freezing/thaw cycles unless specified for special tests.
  • Time windows to analysis vary by test type (e.g.,12–24 h for routine serology; 30 min for ABG; 4 h for CBC/Hb tests).
  • Storage: separate serum/plasma from cells within 2 hours; store sera at appropriate temperatures; some analytes must be transported at specific temperatures.

Post-analytical: reporting and interpretation

  • Reporting includes clinical data, results, reference ranges, and interpretive comments.
  • Ensure correct interpretation in context of reference ranges and patient factors (age, sex, pregnancy, method, reagents).

Units, reference ranges, and interpretation basics

  • Principal units: concentrations reported as mmol/L, µmol/L, nmol/L, pg/mL, mg/L, etc.
  • Reference intervals depend on population and method; most tests assume Gaussian distribution and use Mean ± 2SD for 95% reference range.
    • Reference range: ext{Reference range} = ar{x} \,\pm\, 2\,\sigma
  • Age/gender/population differences and method-dependent differences can shift reference intervals; overlap between healthy and diseased results is common.

Analytical quality: precision, accuracy, and controls

  • Precision definitions:
    • Within-run (intra-assay) and between-run (inter-assay) precision; reported as CV.
    • CV: \text{CV} = \frac{\text{SD}}{\text{mean}} \times 100\%
  • Quality control: use Levey-Jennings charts to track control values over time with ±2SD, ±3SD control limits.
  • Accuracy is reflected in how close results are to true values; calibration and traceability matter.

Diagnostic test performance concepts

  • Sensitivity: \text{Sensitivity} = \frac{TP}{TP+FN} \times 100\%
  • Specificity: \text{Specificity} = \frac{TN}{TN+FP} \times 100\%
  • Predictive values:
    • Positive Predictive Value: \text{PPV} = \frac{TP}{TP+FP} \times 100\%
    • Negative Predictive Value: \text{NPV} = \frac{TN}{TN+FN} \times 100\%
  • Receiver Operating Characteristic (ROC) curves summarize sensitivity vs 1-specificity across cut-offs; the optimal cut-off is often near the top-left corner.
  • Changing the cut-off alters sensitivity and specificity; use population context (screening vs diagnostic testing).

Interference and specimen quality issues

  • Common interferences: hemolysis, icterus, lipemia.
  • Interference indices (limits) example:
    • Hemolysis index (H-Index) up to ~40;
    • Icterus index up to ~60;
    • Lipemia index up to ~150.
  • Drugs and physiological conditions can also cause interference; interpret results with clinical context and in light of potential preanalytical issues.

Practical exam-oriented takeaways

  • Always verify patient identity and sample labeling; use barcodes when available.
  • Remember the dominant role of preanalytical errors in the total error budget (~70–80%).
  • Be able to identify factors that can falsely elevate or depress common tests (e.g., lipemia causing interference in spectrophotometry).
  • Recognize the key concepts of precision (CV), accuracy, reference intervals, and how to interpret them in light of patient factors.
  • Know the basic definitions of sensitivity, specificity, PPV, NPV, and the purpose of ROC analysis.