Table 1. Summary of the results of MGMT promoter methylation testing in cases with unmethylated MGMT promoter according to the reference laboratory. There is a perfect concordance) of MGMT status in all samples irrespective of tissue fixation conditions, testing laboratory, testing technician and time point of testing. Cut-offs for defining MGMT promoter methylated vs. unmethylated cases: for MSRE: 0.32%, for PSQ: 8%. PSQ-% represent the mean-methylation values from four CpG analyzed and calculated from pyrograms.
| ID | Test lab | Test run | Fixation | Method | Technician | Mean value | Result |
|---|---|---|---|---|---|---|---|
| Case 1 | Reference lab | 1 | RCLPE | MSRE | 1 | 0 | unmethylated |
| Case 1 | Sequenom | 2 | RCLPE | Epityper | 1 | 5.33 | unmethylated |
| Case 1 | Test lab 1 | 3 | FFPE | PSQ | 1 | 2.25 | unmethylated |
| Case 1 | Test lab 1 | 4 | FFPE | PSQ | 1 | 1.75 | unmethylated |
| Case 1 | Test lab 1 | 5 | RCLPE | PSQ | 1 | 2.25 | unmethylated |
| Case 1 | Test lab 1 | 6 | RCLPE | PSQ | 2 | 2.75 | unmethylated |
| Case 1 | Test lab 2 | 7 | RCLPE | PSQ | 1 | 2.27 | unmethylated |
| Case 1 | Test lab 2 | 8 | FFPE | PSQ | 1 | 2.55 | unmethylated |
| Case 5 | Reference lab | 1 | RCLPE | MSRE | 1 | 0 | unmethylated |
| Case 5 | Sequenom | 2 | RCLPE | Epityper | 1 | 3.9 | unmethylated |
| Case 5 | Test lab 1 | 3 | FFPE | PSQ | 1 | 1.75 | unmethylated |
| Case 5 | Test lab 1 | 4 | FFPE | PSQ | 1 | 2 | unmethylated |
| Case 5 | Test lab 1 | 5 | FFPE | PSQ | 2 | 1.75 | unmethylated |
| Case 5 | Test lab 1 | 6 | RCLPE | PSQ | 1 | 1.25 | unmethylated |
| Case 5 | Test lab 1 | 7 | RCLPE | PSQ | 2 | 1.5 | unmethylated |
| Case 5 | Test lab 2 | 8 | FFPE | PSQ | 1 | 2.29 | unmethylated |
| Case 5 | Test lab 2 | 9 | RCLPE | PSQ | 1 | 1.27 | unmethylated |
| Case 7 | Reference lab | 1 | RCLPE | MSRE | 1 | 0 | unmethylated |
| Case 7 | Sequenom | 2 | RCLPE | Epityper | 1 | 2.5 | unmethylated |
| Case 7 | Test lab 1 | 3 | FFPE | PSQ | 1 | 3.25 | unmethylated |
| Case 7 | Test lab 1 | 4 | FFPE | PSQ | 1 | 3 | unmethylated |
| Case 7 | Test lab 1 | 5 | FFPE | PSQ | 2 | 3.25 | unmethylated |
| Case 7 | Test lab 1 | 6 | FFPE | PSQ | 2 | 4 | unmethylated |
| Case 7 | Test lab 1 | 7 | RCLPE | PSQ | 1 | 4.25 | unmethylated |
| Case 7 | Test lab 1 | 8 | RCLPE | PSQ | 2 | 3.5 | unmethylated |
| Case 7 | Test lab 2 | 9 | RCLPE | PSQ | 1 | 2.48 | unmethylated |
| Case 7 | Test lab 2 | 10 | FFPE | PSQ | 1 | 2.81 | unmethylated |
| Case 9 | Reference lab | 1 | RCLPE | MSRE | 1 | 0.02 | unmethylated |
| Case 9 | Sequenom | 2 | RCLPE | Epityper | 1 | 1.9 | unmethylated |
| Case 9 | Test lab 1 | 3 | FFPE | PSQ | 1 | 5 | unmethylated |
| Case 9 | Test lab 1 | 4 | FFPE | PSQ | 1 | 5.25 | unmethylated |
| Case 9 | Test lab 1 | 5 | FFPE | PSQ | 2 | 5.25 | unmethylated |
| Case 9 | Test lab 1 | 6 | FFPE | PSQ | 2 | 5.75 | unmethylated |
| Case 9 | Test lab 1 | 7 | RCLPE | PSQ | 1 | 1.5 | unmethylated |
| Case 9 | Test lab 1 | 8 | RCLPE | PSQ | 2 | 1.75 | unmethylated |
| Case 9 | Test lab 2 | 9 | RCLPE | PSQ | 1 | 1.06 | unmethylated |
| Case 9 | Test lab 2 | 10 | FFPE | PSQ | 1 | 3.84 | unmethylated |
FFPE = formalin-fixed and paraffin-embedded; MSRE = methylation-specific restriction enzyme based quantitative polymerase chain reaction; PSQ = pyrosequencing; RCLPE = RCL2-fixed and paraffin-embedded.