Pfluger 2020 [A].

Study characteristics
Patient Sampling	Purpose: Detection of current acute‐phase and current convalescent‐phase SARS‐CoV‐2 infection Design: Two‐group study to estimate sensitivity and specificity [1] Covid patients (n = 75) [2] Pre‐pandemic healthy controls (n = 320) Recruitment: [1] Not stated, hospital inpatients [2] Not stated, retained samples of a pre‐pandemic blood donor cohort Prospective or retrospective: [1] Prospective. First blood sample available after hospitalisation was used [2] Retrospective Sample size: 395 (75) Further detail: [1] COVID‐19 patients (positive RT‐PCR) in March and April of 2020 [2] Retained samples of a pre‐pandemic blood donor cohort collected 01.03.17–09.04.17 Exclusions Not stated
Patient characteristics and setting	Setting: Hospital, inpatient Location: University Medical Center Hamburg‐Eppendorf (UKE), Hamburg, Germany Country: Germany Dates: March and April 2020 Symptoms and severity: Mixed: based on WHO case definitions: critical, 31/75 (41.4%); severe 36/75 (48%); mild 7/75 (9.3%); asymptomatic 1/75 (1.3%) Demographics: Mean age 60.2 ± 15.4, range 16‐93 years; 33.3% female, 66.7% male Exposure history: Not stated. Non‐Covid group 1: Pre‐pandemic controls Source: Pre‐pandemic healthy blood donors, age 18‐70 years (equally distributed), male to female ratio 1:1, collected 01/03/17‐09/04/17 Characteristics: Healthy adults
Index tests	Test name: [A] Anti‐SARS‐CoV‐2 ELISA (IgG) [B] LIAISON SARS‐CoV‐2 S1/S2 IgG [C] Elecsys Anti‐SARS‐CoV‐2 [D] WANTAI SARS‐CoV‐2 Ab ELISA [E] Atellica IM SARS‐CoV‐2 Total (COV2T) Manufacturer: [A] EUROIMMUN AG, Lubeck, Germany [B] DiaSorin S.p.A, Saluggia, Italy [C] Roche Diagnostics Deutschland Gmbh, Mannheim, Germany [D] Beijing Wantai Biological Pharmacy Enterprise Co., Ltd., Beijing, China [E] Siemens Healthcare Gmbh, Erlangen, Germany Antibody: [A] IgG [B] IgG [C] Total Ab [D] Total Ab [E] Total Ab Antigen target: [A] S1‐domain, spike‐protein [B] S1 and S2‐protein [C] N‐protein [D] RBD [E] Spike‐protein Evaluation setting: Laboratory Test method: [A] ELISA [B] CLIA [C] ECLIA [D] ELISA [E] CLIA Timing of samples: Mean time pso was 11.4 days (± 6.6), range 1‐38 days 1‐10 days n = 37 11‐15 days n = 22 16‐38 days pso n = 16 Samples used: plasma/serum Test operator: Laboratory staff Definition of test positivity: [A] > 1.1 ratio (borderline 0.8‐1.1) [B] > 15 AU/mL (borderline 12‐15) [C] > 1 COI [D] > 1 A/C.O (borderline 0.9‐1.1) [E] > 1 Index Blinding reported: Not stated Threshold predefined: Yes (according to manufacturer's instructions)
Target condition and reference standard(s)	Reference standard: Positive RT‐PCR, either by modified E‐gene assay adapted as 'cobas Omni Utility Channel'‐protocol (Ct value < 34 positive in at least 2 independent samples) or by Roche SARS‐CoV‐2 IVD‐Test 9 samples received external ref standard PCR. Samples used: Naso‐pharyngeal swab Timing of reference standard: Not stated Blinded to index test: Yes Incorporated index test: No Definition of non‐COVID cases: Pre‐pandemic Samples used: NA, pre‐pandemic Timing of reference standard: pre‐pandemic Blinded to index test: Yes Incorporated index test: No
Flow and timing	Time interval between index and reference tests: Not stated All patients received same reference standard: No [1] 3 RT‐PCR tests used. 66 samples tested in‐house using 'cobas Omni Utility Channel' or Roche SARS‐CoV‐2 IVD test. 9 samples external RT‐PCR test [2] Pre‐pandemic samples Missing data: Not stated Uninterpretable results: Not stated Indeterminate results: Some samples had borderline results, classed as positive Unit of analysis: Patients
Comparative
Notes	Funding: Partially funded by the BGV (Behorde fur Gesundheit und Verbaucherschutz der Freien und Hansestadt Hamburg). Some authors funded by German Center for Infection Research (DZIF) and some by German Research Foundation (DGF, SFB841) Publication status: Published paper Source: Journal of Clinical Virology Author COI: Marc Lutgehetmann has received travel expenses and speakers' honoraria (Roche Diagnostics, DiaSorin,Biomerieux). Other authors declared no conflict of interest.
Methodological quality
Item	Authors' judgement	Risk of bias	Applicability concerns
DOMAIN 1: Patient Selection
Was a consecutive or random sample of patients enrolled?	Unclear
Was a case‐control design avoided?	No
Did the study avoid inappropriate exclusions?	Unclear
Did the study avoid inappropriate inclusions?	Unclear
Could the selection of patients have introduced bias?		High risk
Are there concerns that the included patients and setting do not match the review question?			High
DOMAIN 2: Index Test (All tests)
DOMAIN 2: Index Test (Antibody tests)
Were the index test results interpreted without knowledge of the results of the reference standard?	Unclear
If a threshold was used, was it pre‐specified?	Yes
Could the conduct or interpretation of the index test have introduced bias?		Unclear risk
Are there concerns that the index test, its conduct, or interpretation differ from the review question?			Low concern
DOMAIN 3: Reference Standard
Is the reference standards likely to correctly classify the target condition?	Yes
Were the reference standard results interpreted without knowledge of the results of the index tests?	Yes
The reference standard does not incorporate the index test	Yes
Could the reference standard, its conduct, or its interpretation have introduced bias?		Low risk
Are there concerns that the target condition as defined by the reference standard does not match the question?			High
DOMAIN 4: Flow and Timing
Was there an appropriate interval between index test and reference standard?	Unclear
Did all patients receive the same reference standard?	No
Were all patients included in the analysis?	Yes
Did all participants receive a reference standard?	Unclear
Were results presented per patient?	Yes
Could the patient flow have introduced bias?		High risk