Homburg, 23.12.05
The aim of the study was to evaluate Medac EBNA-1 IgG in comparison with Biotest EBNA-1-IgG and with a reference in-house IFA (VCA-IgG, VCA-IgM, EBNA-1-IgG). The assays were performed according to the manufacturer’s instructions or to our in-house protocol (Gärtner, Hess et al. 2003).
Interpretation of IFA: ≤ 1:8 negative, 1:16 borderline, ≥ 1:32 positive
The study was divided into five parts
1. 70 EBV negative sera
2. 70 EBV past infections
3. 70 EBV primary infections
4. 50 sera from lung transplant recipients with prior EBV infection
5. 50 sera from patients with rheumatic diseases with prior EBV infection
1. EBV negative sera
Samples: 68 samples from EBV negative patients were selected (two more samples were taken by accident but did not origin from EBV negative individuals, one sample (438707) belonged to a lung transplant recipient and 451116 was a past infection of an immunocompetent individual).
Results: All 68 sera were congruently negative with Medac and Biotest. Five samples showed unspecific results in IFA, the remaining 63 samples were negative.
Conclusion: All samples were correctly identified according to clinical diagnosis
2. Past infections
Samples: 69 samples from patients with past infections were selected (one additional patient aged 68 had a reactivation and was positive in EBV PCR from whole blood with 3000 copies/ml, 458247).
Results: All 69 sera were congruently positive with Medac and Biotest. One sample showed a borderline titer in IFA (1:16), the remaining 68 samples were clearly positive. The sample with the EBV reactivation was EBNA-1 positive in IFA but negative with Medac and Biotest.
Conclusion: All samples were correctly identified according to clinical diagnosis. The fact that the reactivated sample was negative in the EIA is well in accordance with the know fact that EBNA-1 decreases during reactivation.
3. EBV primary infections
Samples: 70 samples from patients with primary EBV infections (all VCA-IgM positive) were analyzed.
Results: 69 sera were congruently negative with Medac and Biotest. One sample was positive in both EIA but negative in IFA (352330). This was a 30 year old immunocompetent patient from an external lab, thus we have no more information on this patient than that a mononucleosis was suspected. Moreover one sample was negative in the EIA but positive in IFA (316758). This was a sample taken two month after primary infection. This might explain the discrepancies. In addition 10 samples had unspecific reactions in EBNA-1-IgG IFA.
Conclusion: 69 from 70 samples were correctly identified according to clinical diagnosis.
4. Sera from lung transplant recipients
Samples: 50 samples from immunocompromised patients after lung transplantation with past EBV infections were selected. Follow-up samples from the same patients were included as indicated by the initials of the name.
Results: 46 sera were congruently positive with Medac and Biotest. Four samples were negative in both assays. Three of them originated from the same patient (226639, 228918, 231265, patient HD); they reacted borderline or positive in IFA. One more sample from another patient was negative with both EIA but positive in IFA (225617, DS). On the other hand 6 out of 7 samples from patient FF reacted negative or borderline in IFA (209600, 212099,212379,215540, 218084, 222767) only one was clearly positive (210236). Moreover, two further patients (HOT 226633 and KB 207666) were negative in IFA but positive in the EIAs. In addition one sample reacted unspecific in IFA.
Conclusion: Since all sera originated from patients with past infections, positive EBNA-1-IgG seem to be correct. Forty-six samples (92%) were positive in EIA. Forty-four samples reacted positive or borderline in IFA (88%).
The lung transplant recipients showed a relevant number of discrepancies. Interestingly, the discrepancies were liked to the individual patient and were not errors by chance.
5. Sera from patients with rheumatic diseases
Samples: 50 samples from different immunocompetent patients with past infections and rheumatic disorders were selected.
Results: 42 samples were congruently positive with Medac and Biotest, 7 were congruently negative and one was discrepant between the two assays (438939).
From the 7 negative sera in both EIA four were concordantly negative in IFA. The remaining discordant three samples are indicated in the table (321463, 417765, 450173 green). On the other hand two further samples reacted negative in IFA but positive in EIA (304324, 418684 red). One additional sample (438939 yellow) was negative in Biotest and IFA but positive using Medac.
Discrepant samples
|
|
Biotest
Cut-off 0.22 |
Medac |
IFA |
Remarks |
|
304342 |
Pos 1.14 |
Pos 30.3 |
8 |
Low positive titer in IFA |
|
418684 |
Pos 1.50 |
Pos 78.2 |
Neg |
|
|
438939 |
Neg 0.11 |
Pos 20,0 |
Neg |
“low positive” in Medac,
“high negative” in Biotest |
|
321463 |
Neg 0.05 |
Neg <9 |
32 |
|
|
417765 |
Neg 0.02 |
Neg <9 |
32 |
|
|
450173 |
Neg 0.05 |
Neg <9 |
<32 |
|
Conclusion: Since all sera originated from patients with past infections, positive EBNA-1-IgG seem to be correct. Using Medac or IFA 43 samples reacted positive (86%), similarly 42 when employing Biotest (84%). However, the patients with rheumatic disorders showed an even higher rate of discrepancies compared to the lung transplant recipients. Four patients were congruently negative in all assays and 6 gave discrepant results.
Summary
The two EIA performed almost identical with 309 identical results out of 310 tested samples (99.7%).
Compared to the clinical diagnosis in seronegative, past and primary infections all but one sample were identified correctly (206 out of 207, 99.5%)
The EIA showed a better performance compared to the reference IFA since a relevant number (16/310, 5.2%) of samples reacted unspecific. In seronegatives, past infections and primary infections 15/207 (7.2%), and in lung transplant recipients and in patients with rheumatic disorders 1/100 (1%) showed unspecific results.
Moreover, the rate of EBNA-1-IgG negative samples in patients with past infections and lung transplantation or rheumatic disorders was 12 (12%) with Medac; 13 (13%) using Biotest and 13 (13%) employing in-house IFA (when borderline titers were included the number rises to 18 (18%) or 19 (19%) when unspecific results were included as well).
Taken together the Medac EBNA-1-IgG showed an excellent performance.
Reference
1. Gärtner, B. C., R. D. Hess, D. Bandt, A. Kruse, A. Rethwilm, K. Roemer and N. Mueller-Lantzsch (2003). "Evaluation of four commercially available Epstein-Barr virus enzyme immunoassays with an immunofluorescence assay as the reference method." Clin Diagn Lab Immunol 10: 78-82.