Favresse 2020a.

Study characteristics
Patient Sampling	Purpose: external validation of a new electrochemiluminescent immunoassay (ECLIA) test that allows the detection of total antibodies 2‐group study to estimate sensitivity and specificity for diagnosis of active disease/identification of previous disease Design: Two groups of samples: [1] patients with a confirmed RT‐PCR SARS‐CoV‐2 diagnosis (n = 97 patients, 140 samples) [2] Non‐SARS‐CoV‐2 sera collected prior to the COVID‐19 pandemic with potential cross‐reactions (n = 79) Recruitment: Retrospective, no further information Prospective or retrospective: Retrospective Sample size: 219 (140) samples, 176 (97) patients Further detail: Not stated
Patient characteristics and setting	Setting: Unclear ‐ probably hospital inpatients because of multiple samples for patients Location: Clinique Saint‐Luc Bouge (SLBO, Namur, Belgium) Country: Belgium Dates: Unclear. "This retrospective study was conducted from May 6 to 12, 2020", but not clear whether these were recruitment dates Symptoms and severity: Not stated Demographics: Not stated Exposure history: Not stated Non‐Covid group 1: [2] Pre‐pandemic controls Source: Between January 2019 and December 2019. Source not stated Characteristics: Potential cross‐reactions (cross‐reactivity test group) were also analysed. Samples in this group included: positive antinuclear antibodies (n = 5), antithyroglobulin antibody (n = 1), anti‐Treponema pallidum antibodies (n = 2), antistreptolysin O (n = 1), antithyroid peroxidase antibodies (n = 4), chikungunya antibody (n = 1), direct Coombs (n = 1), hepatitis B antigen (n = 4), hepatitis C antibodies (n = 7), hepatitis E antibodies (n = 4), HIV antibodies (n = 2), IgA chlamydia pneumoniae (n = 1), IgG chlamydia trachomatis (n = 1), IgG Coxiella burneti (n = 2), IgM Borrelia (n = 1), IgM Coxiella burnetii (n = 1), IgM cytomegalovirus (n = 5), IgM Epstein‐Barr virus viral capsid (n = 5), IgM mycoplasma pneumoniae (n = 6), IgM parvovirus B19 (n = 7), IgM toxoplasma gondii (n = 5), influenza antibodies (n = 6), irregular agglutinins (n = 2), and rheumatoid factor (n = 5).
Index tests	Test name: Elecsys anti‐SARS‐CoV‐2 Manufacturer: Roche Diagnostics Antibody: total antibodies (including IgG) Antigen target: SARS‐ CoV‐2 nucleocapsid Evaluation setting: Laboratory test conducted in the laboratory Test method: electrochemiluminescent immunoassay (ECLIA) Timing of samples: 0‐ ≥ 28 days after positive RT‐PCR test, 0‐6 days post‐PCR+: 45/140 7‐13 days post‐PCR+: 35/140 14‐20 days post‐PCR+: 24/140 21‐27 days post‐PCR+: 15/140 28+ days post‐PCR+: 21/140 0‐ > 28 days after onset of symptoms 0‐6 days pso: 22/129 7‐13 days pso: 28/129 14‐20 days pso: 26/129 21‐27 days pso: 23/129 28+ days pso: 30/129 11 missing data on time pso Samples used: Serum samples Test operator: Laboratory personnel Definition of test positivity: Two thresholds reported: [A] According to the manufacturer, a result < 1.0 is considered negative while a result ≥ 1.0 is considered positive [B] optimal cut‐off provided by ROC curve analyses (i.e. > 0.165) Blinding reported: Not stated Threshold predefined: [A] The test result is given as a cut‐off index (COI). According to the manufacturer, a result < 1.0 is considered negative while a result ≥ 1.0 is considered positive. [B] No for optimised cut‐off
Target condition and reference standard(s)	Reference standard: RT‐PCR performed on the LightCycler® 480 Instrument II using the LightMix® Modular SARS‐CoV‐2 E‐gene set (Roche Diagnostics®) Samples used: respiratory samples (nasopharyngeal swab samples) Timing of reference standard: Not stated Blinded to index test: Yes, prior to index test Incorporated index test: No Definition of non‐COVID cases: Pre‐pandemic Samples used: Not stated Timing of reference standard: Pre‐pandemic Blinded to index test: Yes, prior to index test Incorporated index test: No
Flow and timing	Time interval between index and reference tests: 0‐ ≥ 28 days All patients received same reference standard: No, controls were pre‐pandemic Missing data: Among the 97 patients (140 samples), data about time of symptom onset were available for 92 patients (129 samples). Uninterpretable results: Not stated Indeterminate results: Not stated Unit of analysis: Samples
Comparative
Notes	Funding: Roche Diagnostics provided the kits for the validation. Publication status: Published letter Source: Clinical Chemistry, Volume 66, Issue 8, August 2020, Pages 1104–6 Author COI: J. Douxfils, personal fees from Diagnostica Stago, Roche, Roche Diagnostics, Daiichi‐Sankyo, and Portola, outside the submitted work. J. Douxfils, chief executive officer and founder of QUALIblood sa
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?	No
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?	No
Could the conduct or interpretation of the index test have introduced bias?		High 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?	No
Were results presented per patient?	No
Could the patient flow have introduced bias?		High risk