Suhandynata 2020b [A].

Study characteristics
Patient Sampling	Purpose: Diagnosis of current acute‐phase infection and current convalescent‐phase infection Design: Multi‐group study to estimate sensitivity and specificity [1] Confirmed COVID patients (n = 60) [2] Non‐COVID subjects (n = 179) [2a] Current, other diseases (n = 22) [2b] Current, positive for other antibodies, DNA or IgM/IgG (n = 27) [2c] Current, apparently healthy subjects (n = 20) [2d] Pre‐pandemic samples (n = 110) Recruitment: Not stated Prospective or retrospective: Retrospective Sample size: 239 (60) patients with 339 (160) samples of which 204 (25) were eligible for our review Further detail: Inclusion: [1] Patients which tested PCR‐positive for SARS‐CoV‐2 [2a] Patients which tested PCR‐positive on a respiratory panel nucleic acid (RPNA) test infections other than SARS‐CoV‐2 [2b] Patients which tested positive for antinuclear antibodies (ANA) or anti‐double stranded DNA (dsDNA) or patients with clinically elevated levels of IgM/IgG [2c] Apparently healthy subjects (no respiratory symptoms per self‐report) [2d] Patient samples that had been stored frozen (‐20 degrees C) since 2018 Exclusions not reported
Patient characteristics and setting	Setting: Not stated Location: UC San Diego Health clinical laboratories, California Country: California, USA Dates: Not stated Symptoms and severity: Not stated Demographics: Not stated Exposure history: Not stated Non‐Covid group 1: [2a] and [2b] Cross‐reactivity panel Source: UC San Diego Health clinical laboratories, current (time not stated) Characteristics: Human metapneumovirus n = 4 Influenza A H1‐2009 PCR n = 1 Mycoplasma pneumoniae n = 1 Non‐COVID coronavirus n = 7 Parainfluenza 4 PCR n = 1 Respiratory syncytial virus A n = 2 Respiratory syncytial virus B n = 2 Rhinovirus/enterovirus n = 4 Anti‐dsDNA (> 100 IU/mL) n = 4 Antinuclear antibodies n = 20 Elevated IgG/IgM n = 3 Non‐Covid group 2: [2c] Current, healthy (untested) [2d] Pre‐pandemic Source: [2c] Source not stated, current [2d] UC San Diego Health clinical laboratories, patient samples that had been stored frozen (‐20 degrees C) since 2018 Characteristics: [2c] Apparently healthy subjects (no respiratory symptoms per self‐report) [2d] Not stated
Index tests	Test name: [A] Diazyme DZ‐LITE 2019‐nCoV IgG, IgM (CLIA) Assay Kits (Cat # 130219015M; Cat # 130219016M) [B] Roche Elecsys Anti‐SARS‐CoV‐2 total Ig (Ref # 09203079190) [C] Abbott SARS‐CoV‐2 IgG (Ref # 06R8620) reagent kit Manufacturer: [A] Diazyme [B] Roche [C] Abbott Antibody: [A] IgG, IgM [B] Total antibodies [C] IgG Antigen target: Not stated Evaluation setting: All laboratory tests Test method: [A] CLIA [B] CLIA [C] CMIA Timing of samples: ≤ 7 days post‐PCR+ (n = 43) 8–14 days post‐PCR+ (n = 31) ≥ 15 days post‐PCR+ (n = 25) Samples used: Plasma (Li‐Heparin or K‐EDTA) and serum samples Test operator: Department of Pathology UC San Diego Health Definition of test positivity: [A] Absorbance units per mL (AU/mL), values ≥ 1.00 AU/mL were considered reactive. [B] A cut‐off index (COI; signal of sample/cut‐off); values ≥ 1.00 COI were considered reactive. [C] Index value (S/C); Index values ≥ 1.4 S/C were considered positive. Blinding reported: Not stated Threshold predefined: yes (analysed in a manner consistent with the package inserts)
Target condition and reference standard(s)	Reference standard: Positive for COVID‐19 by a nucleic acid amplification test that had been clinically validated in our laboratory and had an emergency use authorisation (EUA) listing with the US Food and Drug Administration Threshold not stated Samples used: Not stated Timing of reference standard: Not stated Blinded to index test: yes, prior to index test Incorporated index test: no Definition of non‐COVID cases: [2a], [2b] To identify patient specimens containing other PCR‐confirmed microbes, the respiratory pathogen nucleic acid (RPNA) test was performed on the GenMark ePlex. This panel detects Adenovirus (A‐F), coronavirus (229E, HKU1, NL63, OC42), human metapneumovirus, human rhinovirus/enterovirus, influenza A, B and C, influenza 2009 H1N1, parainfluenza (1‐4), respiratory syncytial virus (A and B), chlamydia pneumoniae and mycoplasma pneumoniae. [2c] Untested (no respiratory symptoms per self‐report) [2d] Pre‐pandemic Samples used: [2a] and [2b] Not stated or untested [2c] Untested [2d] Pre‐pandemic Timing of reference standard: Not stated Blinded to index test: yes, prior to index test Incorporated index test: no
Flow and timing	Time interval between index and reference tests: ≤ 7 days post‐PCR+ (n = 43), 8–14 days post‐PCR+ (n = 31), ≥ 15 days post‐PCR+ (n = 25) All patients received same reference standard: no Missing data: 74 COVID samples < 15 days post‐positive PCR not included in review; only 1 sample used per patient per time split (160‐99 = 61 samples excluded from analyses) Uninterpretable results: Not stated Indeterminate results: No borderline range Unit of analysis: [1] Samples but only one sample from each PCR‐positive patient used per specified time frame [2] Patients
Comparative
Notes	Funding: Research Funding: R.T. Suhandynata, Waters Corporation; M.A. Hoffman, Roche Diagnostics The funding organisations played no role in the design of study, choice of enrolled patients, review and interpretation of data, preparation of manuscript, or final approval of manuscript Publication status: Published paper Source: Journal of Applied Laboratory Medicine Author COI: Employment or leadership: None declared Consultant or advisory role: None declared Stock ownership: None declared. Honoraria: None declared Research Funding: R.T. Suhandynata, Waters Corporation; M.A. Hoffman, Roche Diagnostics Expert testimony: None declared Patents: None declared
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?	No
Were the reference standard results interpreted without knowledge of the results of the index tests?	Unclear
The reference standard does not incorporate the index test	Yes
Could the reference standard, its conduct, or its interpretation have introduced bias?		High 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?	Unclear
Did all participants receive a reference standard?	Yes
Were results presented per patient?	Yes
Could the patient flow have introduced bias?		High risk