Lau 2020c.

Study characteristics
Patient Sampling	Purpose: To diagnose current acute‐phase infection or current convalescent‐phase infection Design: Multi‐group study to estimate sensitivity and specificity [1] Confirmed COVID patients, residual leftover sera (n = 353); [2] Non‐COVID Control ‐ [2a] Current healthy healthcare workers (HCWs) (n = 262); [2b] pre‐pandemic samples from our staff health screening (HS) programme in 2018 (n = 718); [2c] Cross‐reactivity panel (229/262 HCW volunteers from [2a] with recent influenza vaccination and 97 samples positive for dengue fever or other antibodies). Group [2a] and parts of [2c] were excluded from our review as they did not have an eligible reference standard. Recruitment: [1] Test samples ‐ Anonymised residual leftover sera (from other routine testing, e.g. renal panels, complete blood count) from subjects who had positive RT‐PCR at Changi General Hospital between April‐June 2020; unclear how recruited [2a] Excluded from review [2b] Stored samples from our staff health screening (HS) programme in 2018, unclear how recruited [2c] Samples of [2a] who had received a flu vaccination within 4 weeks of the antibody test plus samples that tested positive for dengue fever or other antibody‐positive subjects Prospective or retrospective: Both [1] test samples ‐ retrospective [2a] control group ‐ prospective [2b] pre‐pandemic healthy ‐ retrospective [2c] not stated for the 97 additional samples Sample size: 1430 (353) samples of which 1168 (353) were eligible for our review Further detail: Inclusion: [1] Subjects who had positive RT‐PCR at Changi General Hospital between April‐June 2020; [2a] Healthcare workers (HCWs) (laboratory staff, doctors, nurses, and housekeeping staff) volunteers at Changi General Hospital without symptoms of upper respiratory tract infection/fever and two serial antibody testing 14 days apart; [2b] Stored samples from staff health screening (HS) programme in 2018 (Changi General Hospital); [2c] HCW volunteers who had received the latest influenza vaccination (southern hemisphere) within four weeks of their first SARS‐CoV‐2 IgG test (see [2a]) or samples that tested positive for dengue fever or other antibodies [Anti‐HCV, Hepatitis B, anti‐nuclear antibody (ANA), double‐stranded DNA antibody (ds‐DNA), rheumatoid factor (RF), syphilis]. Exclusion: [1] Test group ‐ PCR‐negative samples [2] Not stated
Patient characteristics and setting	Setting: Hospital (Not stated if inpatients only or also outpatients) Location: Changi General Hospital, Singapore Country: Singapore Dates: April‐June 2020 Symptoms and severity: not mentioned (we did not have any data regarding symptom severity in our sensitivity cohort) Demographics: Sensitivity group (n = 279) Age: Mean 50.3 (SD 17.6) range 23 to 98; 234 (83.9%) males, 45 (16.1%) females Exposure history: not mentioned Non‐Covid group 1: [2c] Cross‐reactivity panel Source: [2c] 229/262 from [2a] not eligible for our review 97 additional samples, source and time for additional cross‐reactivity samples not stated Characteristics: [2c] 46 samples that tested positive for dengue fever, 51 other antibody‐positive subjects [Anti‐HCV – 4, Hepatitis B – 29, anti‐nuclear antibody (ANA) – 11, double‐stranded DNA antibody (ds‐DNA) – 1, rheumatoid factor (RF) – 5, syphilis – 1] Non‐Covid group 2: [2b] Pre‐pandemic healthy adults Source: stored samples from staff health screening (HS) programme in 2018 Characteristics: Age: Mean 44.2 (SD 13.4), range 20 to 85; 365 (50.8%) males, 353 (49.2%) females; healthy
Index tests	Test name: Abbott SARS‐CoV‐2‐IgG Manufacturer: Abbott Antibody: IgG Antigen target: Undisclosed epitope on the viral nucleocapsid Evaluation setting: laboratory Test method: qualitative chemiluminescent microparticle immunoassay Timing of samples: 0‐6 days post‐PCR+: 172/279 7‐13 days post‐PCR+: 47/279 14+ days post‐PCR+: 60/279 Samples used: Serum [1] Leftover sera (stored at 4 °C for 10 days) [2a] Serum [2b] Stored serum Test operator: not stated Definition of test positivity: Compared to the mean chemiluminescent signal of a calibrator, an IgG index is derived with a stated cut‐off index (COI) of 1.4 Blinding reported: not stated Threshold predefined: yes by the manufacturer
Target condition and reference standard(s)	Reference standard: RT‐PCR‐ targets the N and E genes using a Qiagen EZ1 extraction system and Rotor Gene Q amplification system. Samples used: Not stated Timing of reference standard: Not stated Blinded to index test: yes, performed prior to index test (74 patients who were not initially suspected of having COVID‐19 but tested positive for SARS‐CoV‐2 RT‐PCR in their subsequent work‐up had samples for antibody test taken prior PCR+ test but these were excluded from analyses). Incorporated index test: No Definition of non‐COVID cases: [2a] NA as excluded from review; [2b] Pre‐pandemic (2018); [2c] Not stated for the additional 97 cross‐reactivity samples. Samples used: [2a] NA as excluded from review; [2b] Pre‐pandemic; [2c} Not stated. Timing of reference standard: [2a] NA as excluded from review; [2b] Pre‐pandemic; [2c] Not stated. Blinded to index test: yes, performed prior to index test Incorporated index test: No for [1], [2b] and eligible 97 samples from [2c]
Flow and timing	Time interval between index and reference tests: [1] 0‐6 days: 172/279; 7‐13 days: 47/279; 14+ days: 60/279; [2b] and remaining [2c] not stated All patients received same reference standard: No [1] rtPCR, [2b] pre‐pandemic, [2c] not stated. Missing data: Out of 353 RT‐PCR samples, 74 were excluded as these inpatients were not initially suspected of having COVID‐19 but tested positive for SARS‐CoV‐2 RT‐PCR in their subsequent work‐up. Of the remaining 279 samples, only 60 were eligible for our review (at least 14 days post‐PCR+). From our review, we also excluded group [2a] and 229 samples of group [2c]. Uninterpretable results: Not stated Indeterminate results: Not stated Unit of analysis: samples ([1] 279 samples from 160 individual SARS‐CoV‐2 RT‐PCR‐positive patients; [2c] patients; [2b] not stated)
Comparative
Notes	Funding: We thank Temasek Holdings Pte Ltd and Abbott Diagnostics, Singapore, for sponsoring the test kits used in this study. Publication status: Published paper Source: Clinica Chimica Acta, Elsevier Author COI: None The authors declared that they had no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
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?	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?	Unclear
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?		Unclear 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?	No
Did all participants receive a reference standard?	Unclear
Were results presented per patient?	No
Could the patient flow have introduced bias?		High risk