Standard Operating Procedure: Amylase and Lipase Testing

1. Purpose and Scope

To measure serum amylase and lipase for the diagnosis and monitoring of acute and chronic pancreatitis, and to aid in the evaluation of hyperamylasemia of non-pancreatic origin.

2. Specimen Requirements

• Type: Serum (plain tube or SST).
• Volume: Minimum 0.5 mL.
• Timing: No special patient preparation; random sampling. For monitoring, consistent timing relative to symptom onset is useful because both enzymes rise and fall over hours to days.
• Interference: Grossly lipemic samples may interfere with turbidimetric lipase methods.
• Stability: Serum is stable 24-48 hours refrigerated; amylase is also stable in urine (spot or 24-hour) for macroamylasemia evaluation when indicated.

3. Safety and Precautions

• Standard PPE (gloves, lab coat, eye protection). All blood samples are potentially infectious.
• Dispose of sharps and biohazard waste per laboratory policy.
• Do not pipette by mouth. Use mechanical pipetting devices.

4. Equipment and Reagents

• Automated chemistry analyzer (spectrophotometric).
• Amylase: Enzymatic colorimetric method using maltotetraose or 2-chloro-4-nitrophenyl maltotrioside (CNP-G3) substrate.
• Lipase: Enzymatic colorimetric method using 1,2-diglyceride or triolein substrate with colipase, coupling to a chromogen at 550-580 nm.
• Urine amylase/creatinine: For clearance ratio studies in macroamylasemia (optional reflex).
• Two levels of quality control material for each assay.

5. Step-by-Step Procedure

1. Centrifuge the clotted specimen at 3000-3500 rpm for 10 minutes.
2. Load serum onto the analyzer per instrument instructions with correct assays and calibration parameters.
3. Run two levels of quality control for amylase and lipase before patient samples.
4. Measure amylase and lipase enzymatically; verify calibration linearity.
5. Dilute and re-run results above the upper analytical limit (common in acute pancreatitis).
6. For isolated elevations of amylase with a normal lipase in the appropriate clinical setting, consider reflex testing: urine amylase, salivary amylase fractionation, or macroamylase screen (amylase:creatinine clearance ratio).
7. Release results after QC acceptance.

6. Quality Control

• Run two levels (normal and abnormal) of control daily, with each calibration, and each new reagent lot.
• Enforce Levey-Jennings rules; investigate trends or shifts before reporting patient results.
• Participate in an external quality assessment (EQA) program for both assays.
• Verify control values fall within acceptable target ranges before patient reporting.

7. Decision limits

The interpretive thresholds below are clinical decision limits; the canonical adult reference intervals are displayed in the Reference Ranges panel.

• Acute pancreatitis: lipase elevation above 3 times the upper limit of normal supports the diagnosis and is more specific than amylase. Lipase rises within 4-8 hours, peaks around 24 hours, and remains elevated longer than amylase, making it the preferred pancreatic enzyme.
• Isolated hyperamylasemia with normal lipase: consider salivary sources (parotitis/mumps, dental surgery), ectopic (fallopian tube rupture, ovarian cyst), macroamylasemia (decreased renal clearance, low urine amylase, normal amylase:creatinine clearance ratio), or renal impairment.
• Renal failure: both amylase and lipase may be modestly elevated without pancreatitis.
• Caveats: lipase is less sensitive in early (first 12 hours) massive pancreatitis; interpretation requires clinical correlation with imaging in atypical presentations.