LoBLoDLoQVerificationChecker

URS & FS — User Requirements and Functional Specification

This document describes the controlled interface and behaviour of LoBLoDLoQVerificationChecker for LoB/LoD/LoQ Verification Checker.

Domain limits and critical parameters

• LoB/LoD/LoQ Verification Checker — Analytical Sensitivity Limits Verification per CLSI EP17-A3
• ℹ️ Utility performs statistical verification of sensitivity limits according to CLSI EP17-A3, FDA Bioanalytical Method Validation Guidance and ICH Q2(R2):
• LIMIT OF BLANK (LoB): Upper measurement limit for analyte-free samples. Parametric: Mean_blank + 1.645×SD_blank.
• LIMIT OF DETECTION (LoD): Minimum concentration distinguishable from zero. Parametric: LoB + 1.645×SD_low. Must be > LoB.
• LIMIT OF QUANTITATION (LoQ): Minimum concentration measurable with acceptable precision and accuracy. CV ≤ target% AND |Bias| ≤ target%.
• DATA SUFFICIENCY (EP17-A3): ≥60 blank measurements, ≥60 low-level measurements, ≥20 LoQ replicates.
• • LoD ≤ LoB = logically impossible. Calculation or data error.
• • Insufficient measurements make limits statistically unjustified.
• • Method transfer requires VERIFICATION, not just copying of limits.
• ⚠️ CRITICAL:
• • Blank/Low-Level Measurements ≥ 60 each
• • LoD > LoB (mandatory logical check)
• • LoQ CV ≤ Target% | LoQ |Bias| ≤ Target%
• • Empirical vs Declared Difference ≤ Max allowed %
• • LoQ Replicates ≥ 20
• Key features: Three-limit verification, EP17-A3 data sufficiency check, LoQ precision/accuracy assessment, logical consistency validation, four-tier classification, CLSI/FDA/ICH compliance.
• 💡 Tips: Plan ≥60 blanks + ≥60 low-level BEFORE starting. Use non-parametric if non-normal. Set LoQ targets from clinical requirements. Retain all raw data in validation dossier.
• ⚠️ Note: Performs STATISTICAL VERIFICATION from summary data. For initial establishment, use specialized software (MedCalc, Analyse-it, R 'MDL') with raw measurements. This utility is for independent verification of pre-calculated limits.

URS — User Requirements Specification

input.csv contract

AssayName,Analyte,VerificationType,Method,NumBlankSamples,NumBlankReplicates,BlankMean,BlankSD,DeclaredLoB,NumLowSamples,NumLowReplicates,LowSampleMean,LowSampleSD,DeclaredLoD,TargetAlpha,TargetBeta,DeclaredLoQ,LoQ_CV_Observed,MaxLoQ_CV_Percent,LoQ_Bias_Observed_Percent,MaxLoQ_Bias_Percent,NumLoQReplicates,MinLoQReplicates,EmpiricalLoB,EmpiricalLoD,EmpiricalLoQ,MaxLoB_Diff_Percent,MaxLoD_Diff_Percent,MaxLoQ_Diff_Percent
OncoLiquid_cfDNA,EGFR_L858R_VAF,Verification,Parametric,10,6,0.0001,0.00008,0.0003,6,10,0.0012,0.00035,0.0008,0.05,0.05,0.002,15.5,20.0,8.2,20.0,25,20,0.00028,0.00075,0.0019,20,25,25
MRD_Panel_v2,TP53_ctDNA,Transfer,NonParametric,4,5,0.0005,0.0003,0.0012,3,5,0.003,0.0015,0.005,0.05,0.05,0.008,28.0,20.0,22.0,20.0,15,20,0.0011,0.0032,0.012,20,25,25

Input validation rules

FS — Functional Specification

Example output.json

{
  "utilityId": "loblodloqverificationchecker",
  "utilityFolder": "LoBLoDLoQVerificationChecker",
  "package": "LiquidBiopsy",
  "overallStatus": "PASS|WARNING|FAIL",
  "sourceFile": "input.csv",
  "processedAtUtc": "2026-06-10T00:00:00Z",
  "checks": [
    {
      "parameter": "AssayName",
      "value": "OncoLiquid_cfDNA",
      "status": "PASS|WARNING|FAIL",
      "message": "Deterministic rule-based check result",
      "ruleReference": "FS-RULE-001"
    },
    {
      "parameter": "Analyte",
      "value": "EGFR_L858R_VAF",
      "status": "PASS|WARNING|FAIL",
      "message": "Deterministic rule-based check result",
      "ruleReference": "FS-RULE-002"
    },
    {
      "parameter": "VerificationType",
      "value": "Verification",
      "status": "PASS|WARNING|FAIL",
      "message": "Deterministic rule-based check result",
      "ruleReference": "FS-RULE-003"
    },
    {
      "parameter": "Method",
      "value": "Parametric",
      "status": "PASS|WARNING|FAIL",
      "message": "Deterministic rule-based check result",
      "ruleReference": "FS-RULE-004"
    },
    {
      "parameter": "NumBlankSamples",
      "value": "10",
      "status": "PASS|WARNING|FAIL",
      "message": "Deterministic rule-based check result",
      "ruleReference": "FS-RULE-005"
    },
    {
      "parameter": "NumBlankReplicates",
      "value": "6",
      "status": "PASS|WARNING|FAIL",
      "message": "Deterministic rule-based check result",
      "ruleReference": "FS-RULE-006"
    },
    {
      "parameter": "BlankMean",
      "value": "0.0001",
      "status": "PASS|WARNING|FAIL",
      "message": "Deterministic rule-based check result",
      "ruleReference": "FS-RULE-007"
    },
    {
      "parameter": "BlankSD",
      "value": "0.00008",
      "status": "PASS|WARNING|FAIL",
      "message": "Deterministic rule-based check result",
      "ruleReference": "FS-RULE-008"
    },
    {
      "parameter": "DeclaredLoB",
      "value": "0.0003",
      "status": "PASS|WARNING|FAIL",
      "message": "Deterministic rule-based check result",
      "ruleReference": "FS-RULE-009"
    },
    {
      "parameter": "NumLowSamples",
      "value": "6",
      "status": "PASS|WARNING|FAIL",
      "message": "Deterministic rule-based check result",
      "ruleReference": "FS-RULE-010"
    },
    {
      "parameter": "NumLowReplicates",
      "value": "10",
      "status": "PASS|WARNING|FAIL",
      "message": "Deterministic rule-based check result",
      "ruleReference": "FS-RULE-011"
    },
    {
      "parameter": "LowSampleMean",
      "value": "0.0012",
      "status": "PASS|WARNING|FAIL",
      "message": "Deterministic rule-based check result",
      "ruleReference": "FS-RULE-012"
    }
  ],
  "criticalFindings": [],
  "warnings": [],
  "audit": {
    "inputHash": "sha256:<calculated at runtime>",
    "rulesVersion": "<utility executable version>",
    "documentation": "LoBLoDLoQVerificationChecker.documentation.html"
  }
}

Traceability matrix

IQ/OQ/PQ test scenarios

QA/QC and change control

• Do not rename columns without updating validator, documentation and test set.
• Retain input.csv, output.json, executable version and checksum.
• Before production use, perform IQ/OQ/PQ or equivalent CSV/CSA verification.
• Critical limits shall be verified against the approved specification, registration dossier and local SOPs.
• The utility provides structured QC decision support; final release decision remains with QA/QP and approved procedures.