Multiplex RT-PCR-based detections of CEA, CK20 and EGFR in colorectal cancer patients

Figure 1 Detection of carcinoembryonic antigen, cytokeratin 20 and epidermal growth factor receptor mRNAs in colorectal cancer tissue samples, HT-29 colorectal cancer cells and colorectal cancer patient peripheral blood samples. A: Detection of carcinoembryonic antigen (CEA), cytokeratin 20 (CK20) and epidermal growth factor receptor (EGFR) mRNA in pairs of colorectal cancer (CRC) and normal adjacent tissue samples. Lane 0 = molecular weight marker (100 bp); lane 1 = normal tissue; lane 2 = CRC tissue; B: Assessment of the sensitivity of the reverse transcription polymerase chain reaction (RT-PCR) detections for CEA, CK20 and EGFR mRNA. Lane 0 = molecular weight marker (100 bp); lane 1 = 10⁵ HT-29 cells in 3 mL of normal blood; lane 2 = 10⁴ HT-29 cells in 3 mL of normal blood; lane 3 = 10³ HT-29 cells in 3 mL of normal blood; lane 4 = 10² HT-29 cells in 3 mL of normal blood; lane 5 = 10 HT-29 cells in 3 mL of normal blood; lane 6 = 1 HT-29 cell in 3 mL of normal blood; lane 7 = normal blood; C: Assessment of the sensitivity of the multiplex RT-PCR detections for CEA (131 bp), CK20 (371 bp) and EGFR (474 bp) mRNA. Lane 0 = molecular weight marker (100 bp); lane 1 = 10⁵ HT-29 cells in 3 mL of normal blood; lane 2 = 10⁴ HT-29 cells in 3 mL of normal blood; lane 3 = 10³ HT-29 cells in 3 mL of normal blood; lane 4 = 10² HT-29 cells in 3 mL of normal blood; lane 5 = 10 HT-29 cells in 3 mL of normal blood; lane 6 = 1 HT-29 cell in 3 mL of normal blood; lane 7 = normal blood; D: An example of multiplex RT-PCR-based detection pattern of tumor marker transcripts in the peripheral blood samples of CRC patients. Left: lane 0 = molecular weight marker (100 bp); lane 1 = patient sample positive for CEA (131 bp) and CK20 (371 bp) mRNA; lane 2 = patient sample positive for CEA (131 bp) mRNA; lane 3 = patient sample positive for CEA (131 bp), CK20 (371 bp) and EGFR (474 bp) mRNA; lane 4 = patient sample negative for all markers. Right: lane 0 = molecular weight marker (100 bp); lanes 1-4 = amplification of β-actin (172 bp) cDNA of the patient samples presented in the left side of this panel.

Figure 2 Kaplan-Meier analysis comparing the overall survival of patient groups as developed according to transcript detection patterns for carcinoembryonic antigen, cytokeratin 20 and epidermal growth factor receptor. Group 1: Carcinoembryonic antigen (CEA) positive; Group 2: Positive for 2 markers (CEA and cytokeratin 20); Group 3: Positive for all markers; Group 4: Combination of groups 1 and 2; Group 5: Combination of groups 2 and 3. Log Rank test P values were as follows: Groups 1 vs 3: P = 0.002; Groups 2 vs 3: P = 0.007; Groups 4 vs 3: P = 0.001; Groups 1 vs 5: P = 0.041; Groups 1 vs 2: P = 0.116.