Troubleshooting / Common Issues
Calculated LoB exceeds manufacturer's claim
Possible causes
- –Blank measurement distribution in your laboratory is wider or higher than the manufacturer's reference population
- –Reagent or calibrator stability issue — older or improperly stored reagents can increase background signal
- –Matrix interference from the blank sample material used in the study
- –Instrument cleanliness or carry-over from previous high-concentration measurements
Recommended action
Inspect the blank sample distribution histogram for unusually elevated values or wide spread. Verify reagent storage conditions and expiration dates. Check instrument maintenance records — carry-over from high-concentration specimens can artificially elevate blank readings. If the issue is isolated to one reagent lot, consider whether that lot is an outlier or whether additional lots should be tested.
Large difference in LoB between reagent lots
Possible causes
- –Genuine lot-to-lot variability in reagent non-specific signal
- –One lot stored or handled differently, causing degradation
- –Different blank sample pools used with each lot
Recommended action
Review per-lot statistics in the Analysis by Reagent Lot section. If one lot's blank mean or SD is substantially different from the others, investigate that lot's storage and handling history. Testing an additional lot can help determine whether the outlying lot is anomalous or whether variability is inherent to the reagent system.
Blank distribution appears bimodal or heavily skewed on the histogram
Possible causes
- –Data from two reagent lots or days with very different blank levels are plotted together
- –Some samples labeled 'Blank' may contain trace analyte and should be reclassified
- –Instrument carry-over from high-concentration samples run prior to blank measurements
Recommended action
Review per-lot histograms in the Analysis by Reagent Lot section to determine whether the bimodality is driven by lot differences. Verify the sample type classification in the raw data. If carry-over is suspected, review the measurement sequence and instrument maintenance records.
Insufficient total measurements — study does not meet minimum requirements
Possible causes
- –Fewer than 5 unique blank or low-level samples enrolled
- –Fewer than 4 replicates per sample per reagent-day combination
- –Fewer than 3 test days
- –Fewer than 2 reagent lots
Recommended action
Review CLSI EP17-A2 minimum requirements: 5 blank samples, 5 low-level samples, 3 days, 4 replicates per sample per reagent-day combination, and 2 reagent lots — yielding a minimum of 60 blank measurements per lot. Collect additional measurements to meet these minimums before finalizing the LoB result, as underpowered studies may produce unreliable estimates.