CSM Lab Medicine Branch · 2025 · Expert Consensus

Blood cfDNA Methylation Tumor Markers — Laboratory & Clinical Consensus 2025

血液游离 DNA 甲基化肿瘤标志物实验室检测与临床应用专家共识（2025 版）
11 consensus statements · PCR-focused (all NMPA-approved are PCR) · lab workflow → clinical application · adjunct diagnosis mature, screening cautious
Chinese Society of Laboratory Medicine
Chin J Lab Med 2026;49(2):160–171 · GRADE evidence · 33 experts
11
consensus statements
PCR
focused platform
Lab→Clinic
full pathway
Adjunct
diagnosis mature

1Background & Framework

33
expert panel
4
GRADE levels
2025.8
evidence cutoff
  • cfDNA: 150–200 bp fragments from apoptosis/necrosis — liquid biopsy core.
  • Advantages: non-invasive · sensitive · traceable · dynamic monitoring.
  • Scope: all NMPA-approved blood cfDNA methylation assays are PCR-based — consensus focuses on PCR.

2Laboratory Detection Essentials

Plasma
preferred (no serum)
EDTA
anticoagulant
5–10 ml
blood volume
  • Sample: cfDNA-stabilized tube (Streck); gentle invert 8–10×; two-step centrifugation; never freeze blood.
  • Extraction: magnetic bead + automation — most used for batch testing.
  • Conversion: bisulfite — accurate, repeatable, recommended.
  • QC: performance validation (accuracy · precision · LOD) before clinical use.
8–10×
gentle inversion
2-step
centrifugation
4 zones
lab separation
  • Lab layout: reagent prep · specimen prep · amplification · product analysis — physically independent.
  • ISO15189: quality management; gene-amplification regulation compliance.
  • Calibration: NMPA instruments/consumables; annual calibration; batch QC of reagents.
  • Hemolysis/icterus/lipemia: re-collect samples.
Certified
personnel training
6-mo
capability assessment
≥1 yr
reporting experience
PCR / dPCR
clinical
Nucleic MS
mid
NGS
research
Personnel: certified gene-amplification training + methylation-specific training; capability assessment every 6 months.

3Laboratory Standards — Consensus 1–8

1
Lab qualification: licensed gene-amplification lab; separated zones; avoid contamination.
强推
2
Quality system: pre/analytical/post SOPs; full performance validation.
强推
3
Personnel: certified training; reporting ≥ intermediate title + 1-yr experience.
强推
4
Plasma sampling: cfDNA tube; no freeze; two-step centrifugation.
强推
5
Extraction: magnetic bead + automation recommended.
强推
6
Conversion: bisulfite — accurate & repeatable.
强推
7
Methods: PCR/dPCR clinical; NGS screening; mass-spec mid-throughput.
强推
8
Interpretation: QC-passed results; clinical consultation.
强推

4Clinical Applications — Consensus 9–11

Adjunct diagnosis (Rec 9)
Recommended for CRC · liver · gastric · esophageal · lung — high specificity.
Screening (Rec 10)
Not yet alone for asymptomatic — low precancer sensitivity.
Prognosis/MRD (Rec 11)
Promising — MRD · drug guidance · tracing; evidence accumulating.

5Evidence Highlights

82.7/96.9
Septin9/SDC2/BCAT1 Se/Sp
17.5×
MRD recurrence risk
0.88
EpiPanGI Dx AUC
96.9%
Sp · triple combo
86.4%
ColonSecure Se
82.8/95.8
GutSeer Se/Sp
  • Adjunct: Septin9 Sp >90%, Se 70–80% (dPCR >80%); liver best (AFP+US joint Se 87.2%).
  • Screening: Septin9 Se 48.2% (advanced adenoma 11.2%) — insufficient for precancer; MCED models (Galleri Se 51.5/Sp 99.5; PanSeer 95.7%).
  • MRD: Septin9/BCAN/BCAT1/IKZF1/VAV3 — postoperative (+) vs (−) 17.5× recurrence risk.
  • dPCR edge: 4-marker dPCR for NSCLC reaches Se 90% — outperforms regular PCR.
  • Quality link: interpretation only after QC pass; lab provides clinical consultation.
  • Future: AI/ML multi-marker models · standardization · population heterogeneity research.
  • Lab challenge: cfDNA low concentration · short half-life · high background — standardized QC essential.
  • Positioning: adjunct diagnosis first; screening & MRD advancing with MCED & multi-marker AI models.
MCED: cfDNA methylation is the core marker (2025 liquid-biopsy multi-cancer consensus); UK NHS RCT with Galleri (>140k).
Clinical Significance