Purpose

Define user requirements for a standalone FUZKK utility that accepts laboratory CSV data, evaluates the records using limits embedded in code, and produces LabWare-compatible JSON.

Scope

The utility is intended for preliminary QC/QA review, integration testing, LIMS/LabWare flow and evidence-trail preparation. Final release decisions remain under the laboratory's validated procedure and responsible personnel.

Users

QC analyst, QA reviewer, CSV/validation engineer, LIMS/LabWare integration engineer, responsible laboratory specialist.

User requirements

1. The utility shall run without arguments and print its self-description, a built-in input.csv example from GetDemoData(), and demo evaluation for the embedded records.
2. The utility shall run with two arguments: input.csv output.json.
3. The utility shall not read input.csv and shall not write output.json when started without arguments.
4. CSV numeric values shall be parsed using CultureInfo.InvariantCulture.
5. Output shall be generated as LabWare-compatible JSON with Header, Samples, Results, Status, StatusCode, ErrorMessage, Description and DescriptionEN.
6. For PASS records, ErrorMessage shall be an empty string.
7. Embedded limits shall follow this priority: Ph. Eur. → British Pharmacopoeia / UK implementation → EAEU / regional requirements → EMA/ICH/EU guidance → USP fallback.
8. If an exact monograph is not known, strict standard API limits are used where applicable: assay 98–102%, total impurities ≤1.0%, individual impurity ≤0.5%.
9. For biologics and mAb-like products, aggregation, sterility and endotoxin checks shall be included where relevant to the utility purpose.
10. If a parameter may arrive in different units, the unit shall be represented as a separate input field or explicitly reflected in the input.csv field name.

Input CSV

BatchNumber,ADCName,Unconjugated_Ab_Percent,Half_Conjugated_Percent,Fully_Conjugated_Percent,Clearance_Rate_mL_day_kg,Target_Occupancy_Percent,ProcessYield_Percent,ConjugationMethod
ADC-UAB-2026-001,Trastuzumab-Deruxtecan,2.5,10.0,87.5,12.0,92.0,85.0,Site-Specific
ADC-UAB-2026-002,Sacituzumab-Govitecan,4.0,12.0,84.0,14.5,85.0,78.0,Random
ADC-UAB-2026-003,Polatuzumab-Vedotin,1.0,5.0,94.0,10.0,95.0,90.0,Site-Specific

input.csv fields

Utility description

ADC Unconjugated Antibody Tracker — ADC Unconjugated Antibody Tracker

ADC Unconjugated Antibody Tracker — ADC Unconjugated Antibody Tracker

ℹ️ Utility controls content of unconjugated and partially conjugated antibody:
• Unconjugated antibody DAR0 (≤5.0%)
• Partially conjugated forms (≤15.0%)
• Target occupancy (≥80%)
• Process yield (≥70%)

⚠️ CRITICAL: Unconjugated antibody competes for target without delivering toxin!
This reduces therapeutic efficacy and increases dose.

Usage:
ADCUnconjugatedAntibodyTracker.exe → demo mode (console output)
ADCUnconjugatedAntibodyTracker.exe input.csv output.json → evaluate your data

Input format:
BatchNumber,ADCName,Unconjugated_Ab_Percent,Half_Conjugated_Percent,Fully_Conjugated_Percent,Clearance_Rate_mL_day_kg,Target_Occupancy_Percent,ProcessYield_Percent,ConjugationMethod

Example:
ADC-UAB-2026-001,Trastuzumab-Deruxtecan,2.5,10.0,87.5,12.0,92.0,85.0,Site-Specific

— WHY IS THIS NEEDED?
Unconjugated antibody control is critical for ADC efficacy:
• Unconjugated antibody (DAR0) binds to tumor antigen but does not kill cell
• It blocks access for full ADC molecules, reducing overall therapy efficacy
• Partially conjugated forms may have unstable pharmacokinetics
• Removal of unconjugated antibody (purification) is complex and may reduce product yield
• For site-specific conjugation, DAR0 level is typically lower than for random

⚠️ CRITICAL:
• Unconjugated antibody ≤5.0% — ensures high target occupancy by active forms
• Partially conjugated forms ≤15.0% — product homogeneity control
• Target occupancy ≥80% — guarantees sufficient toxin delivery to tumor
• Process yield ≥70% — economic viability of production
• High DAR0 level requires increased drug dose, increasing side effect risk

Key features:
• Assessment of DAR0 impact on receptor occupancy (Target Occupancy)
• Control of intermediate conjugation forms
• Monitoring of purification process yield from unconjugated antibody
• Comparison of conjugation methods (Random vs Site-specific)

Critical parameters:
• Unconjugated antibody (DAR0): ≤5.0%
• Partially conjugated forms: ≤15.0%
• Target occupancy: ≥80%
• Process yield: ≥70%

💡 Usage tips:
1. Use HIC-HPLC to separate forms by hydrophobicity (DAR)
2. Site-specific conjugation (e.g., via cysteines or glycans) gives narrower DAR0 profile
3. Optimize chromatography conditions for maximum DAR0 removal without product loss
4. Control target occupancy in in vivo models to confirm efficacy
5. If high DAR0 level observed, review conjugation reaction stoichiometry

⚠️ Note: Unlike free toxin, unconjugated antibody is not toxic itself, but it "steals" targets from the drug. This phenomenon is called competitive inhibition. For high-antigen-expressing tumors this is less critical, but for low-expressing targets DAR0 level must be minimal.

Traceability and limitations

• The URS is used as the source document for functional specification, CSV review and later validation work.
• This document does not replace an approved pharmacopoeial monograph, validated analytical method or internal product specification.
• For product-specific limits, the approved customer specification takes priority.

The functional specification describes the behaviour of the standalone FUZKK console utility, input-data format, evaluation algorithm and output JSON structure.

Functional flow

1. Main() checks the number of arguments.
2. If no arguments are provided: PrintHello() prints the description and built-in input.csv example, then RunDemoEvaluation() executes Evaluate() over GetDemoData() and prints demo JSON.
3. If two arguments are provided: RunWithFiles(input.csv, output.json) reads CSV, evaluates each record and writes LabWare-compatible JSON.
4. LoadData() uses CultureInfo.InvariantCulture and shall not be called in no-arguments mode.
5. Evaluate() returns a named tuple with BatchNumber, ProductName, Parameters, CriticalFailCount, WarningCount, Recommendation and RecommendationEN.
6. GetIssues() builds messages for ErrorMessage in WARNING/FAIL cases.

Evaluation rules

• PASS: CriticalFailCount = 0 and WarningCount = 0.
• WARNING: CriticalFailCount = 0 and WarningCount > 0.
• FAIL: CriticalFailCount > 0.
• ERROR: exception during reading or processing.
• ErrorMessage remains empty for PASS.
• Limits are embedded in Program.cs; no external limit configuration is required.

Input and fields

BatchNumber,ADCName,Unconjugated_Ab_Percent,Half_Conjugated_Percent,Fully_Conjugated_Percent,Clearance_Rate_mL_day_kg,Target_Occupancy_Percent,ProcessYield_Percent,ConjugationMethod
ADC-UAB-2026-001,Trastuzumab-Deruxtecan,2.5,10.0,87.5,12.0,92.0,85.0,Site-Specific
ADC-UAB-2026-002,Sacituzumab-Govitecan,4.0,12.0,84.0,14.5,85.0,78.0,Random
ADC-UAB-2026-003,Polatuzumab-Vedotin,1.0,5.0,94.0,10.0,95.0,90.0,Site-Specific

Output JSON

{
  "Header": {
    "UtilityName": "ADC_Unconjugated_Antibody_Tracker",
    "Version": "1.0.0",
    "Timestamp": "UTC",
    "InstrumentID": "FUZKK-QC-WORKSTATION",
    "OperatorID": "Admin"
  },
  "Samples": [
    {
      "SampleID": "from BatchNumber",
      "BatchNumber": "from CSV",
      "ProductName": "from CSV",
      "TestName": "utility-specific test",
      "AnalysisCode": "utility-specific code",
      "Status": "PASS | WARNING | FAIL | ERROR",
      "StatusCode": "1 | 2 | 0 | -1",
      "ErrorMessage": "",
      "Description": "Russian recommendation",
      "DescriptionEN": "English recommendation",
      "Results": [
        {
          "ParameterName": "parameter",
          "ResultValue": 0.0,
          "UnitOfMeasure": "unit",
          "SpecificationLimit": "limit",
          "IsWithinSpec": true
        }
      ]
    }
  ]
}