Charlton 2020 [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 (28 patients, 46 samples) [2] Pre‐pandemic non‐COVID (50 samples) [3] Cross‐reactivity non‐COVID samples [62 samples: pre‐pandemic (n = 15) and concurrent (n = 47)] Recruitment: [1] Hospitalised (or ambulatory) patients confirmed to be positive for SARS‐CoV‐2 upon nasopharyngeal swab or endotracheal aspirate testing by rRT‐PCR [2] Negative samples were retrieved from bio‐banked sera stored at the public health laboratory (Alberta Precision Laboratories) in Alberta collected before 1 November 2019. [3] Convalescent phase sera (either retrieved from stored sera or prospectively collected) Prospective or retrospective: [1] Unclear [2] Retrospective [3] Prospective and retrospective Sample size: 158 (46) samples Further detail: [1] Patients who tested positive for SARS‐CoV‐2 by rRT‐PCR [2] Serum samples stored prior to 1 November 2019 [3] Not stated
Patient characteristics and setting	Setting: Hospital inpatients (26/28; 93%) and ambulatory (2/28; 7%) Location: Not stated [history taken by Alberta Health Services Communicable Diseases Team (Public Health)] Country: Alberta, Canada Dates: Not stated Symptoms and severity: 2/28 (7%) ambulatory 26/28 (93%) hospitalised 9/28 ICU 7/28 Need for mechanical ventilation 1/28 Pulmonary embolism 26/28 COVID pneumonia 1/28 No COVID pneumonia 1/28 Unknown 13/28 acute respiratory distress syndrome 14/28 no acute respiratory distress syndrome 1/28 unknown Demographics: Mean age of patients was 70.1 (median 73; range, 34 to 102 years), 12/28 (43%) female Exposure history: Travel‐related exposures ‐ yes 4 (14%) (USA n = 2; United Arab Emirates n = 1; within Canada n = 1) ‐ no 23 (82%) ‐ unknown 1 (4%) Contact with traveller ‐ yes 6 (21%) ‐ no 21 (75%) ‐ unknown 1 (4%) Infection related to outbreak in long‐term‐care/continuing care facility 9 (32%) Non‐Covid group 1: [2] Pre‐pandemic controls Source: Bio‐banked sera stored at the public health laboratory (Alberta Precision Laboratories) in Alberta collected before 1 November 2019. Characteristics: Not stated Non‐Covid group 2: [3] Cross‐reactivity samples Source: 15 sera were collected prior to the first case of SARS‐CoV‐2 diagnosis in Alberta, and 47 were collected after the first case of SARS‐CoV‐2 was detected in Alberta. Characteristics: The sera were from patients who had tested negative for COVID‐19 by in‐house rRT‐PCR but positive for other viruses as follows (with the number of sera used): ‐ influenza A virus (n = 5), ‐ influenza B virus (n = 5), ‐ respiratory syncytial virus (RSVA, n = 6; RSVB, n = 1), ‐ rhinovirus/enterovirus (n = 6), ‐ human metapneumovirus (HMPV; n = 5), ‐ parainfluenza virus (PIV‐1 and PIV‐4; n = 4), ‐ CoV‐229E (n = 6), ‐ CoV‐NL63 (n = 11), ‐ CoV‐OC43 (n = 7), or ‐ CoV‐HKU1 (n = 7). One patient was positive for multiple viruses (RSVA and enterovirus/rhinovirus).
Index tests	Test name: [A] SARS‐CoV‐2 IgG assay [B] EDI novel coronavirus COVID‐19 IgM and IgG ELISA [C] a novel coronavirus COVID‐19 IgM and IgG assay [D] SARS‐CoV‐2 S1/S2 IgG [E] anti‐SARS‐CoV‐2 ELISA IgA and IgG assay [F] anti‐SARS‐CoV‐2 [G] Rapid Response [H] 2019 nCoV IgM/IgG detection kit [I] SARS‐CoV‐2 IgG/IgM Ab test kit [J] Novel coronavirus IgG/IgM test kit [K] One Step Test for novel coronavirus [L] 2019‐nCoV Ab test Manufacturer: [A] Abbott Laboratories, Abbott Park, IL, USA [B] Epitope Diagnostics Inc., supplied by Affinity Diagnostics Corp., Toronto, ON, Canada [C] DRG International Inc., supplied by Bio‐Rad, Hercules, CA, USA [D] DiaSorin, Stillwater, MN, USA [E] Euroimmun, Mississauga, ON, Canada [F] Roche Diagnostics, Indianapolis, IN, USA [G] BTNX, Markham, Ontario, Canada [H] Biolidics Limited, Singapore [I] Anhui Deep Blue Medical Technology Co., Ltd., Anhui, China [J] Genrui; Genrui Biotech Inc., Shenzhen, China [K] Getein Biotech Inc., Nanjing, China [L] Innovita Biological Technology Co. Ltd., Qian’an, Hebei, China Antibody: [A] IgG [B] IgM and IgG [C] IgM and IgG [D] IgG [E] IgA and IgG [F] Total antibodies (including IgG) [G]‐[L] IgM and IgG Antigen target: [A] Recombinant antigen nucleocapsid protein [B] Recombinant antigens of the RBD and spike‐protein [C] Antibodies recognising recombinant nucleocapsid proteins and peptides [D] IgG antibodies directed against the S1 and S2 domains of the spike‐protein [E] Recombinant S1 domain of the structural protein [F] Recombinant protein representing the nucleocapsid antigen [G], [I], [J], [L] Target unspecified [H] Recombinant protein, target unspecified [K] Recombinant nucleocapsid and spike proteins Evaluation setting: [A]‐[F] Lab test used in lab [G]‐[L] POCT used in lab Test method: [A] chemiluminescent microparticle immunoassay [CMIA] [B] ELISA [C] ELISA [D] chemiluminescence immunoassay [CLIA] [E] ELISA [F] electrochemiluminescence immunoassay (ECLIA) [G]‐[L] Lateral flow test Timing of samples: [A]‐[L] 0‐14 days pso 21/42 15‐21 days pso 11/42 > 21 days pso 10/42 Samples used: [A]‐[L] Serum (all kits assessed using same patient samples from single‐use aliquots). Samples collected, spun down (3000 rpm for 10 min), aliquoted into single‐use aliquots, and frozen at ‐80°C until the time of testing [G]‐[L] Cross‐reactivity panel [3] was not assessed on the POCTs. Test operator: [A]‐[L] Lab personnel Results read independently by two laboratorians; in case of discrepancy, a third laboratorian reading was used as an arbitrator (+/‐/‐ was considered equivocal, +/‐/+ was considered positive). Definition of test positivity: [A]‐[F] as per manufacturer specifications using cut‐offs as described in the package inserts. All values greater than the published cut‐off were considered positive. [G]‐[L] any banding detected for either IgM or IgG. Faint banding was considered positive. Assays where the control line was absent were considered invalid. Testing was performed as per manufacturer specifications. Blinding reported: not stated Threshold predefined: [A]‐[F] yes as per manufacturer specifications [G]‐[L] Yes, visual‐based
Target condition and reference standard(s)	Reference standard: rRT‐PCR, threshold not reported Samples used: Nasopharyngeal swab (27/28) or endotracheal aspirate (1/28) Timing of reference standard: All dates of symptom onset were reported earlier than the date of diagnostic sample collection (mean, 16 days [range, 2 to 48 days]). Blinded to index test: yes, done prior Incorporated index test: no Definition of non‐COVID cases: [2] Pre‐pandemic [3] Pre‐pandemic or in‐house rRT‐PCR on nasopharyngeal swab testing Samples used: [2] Pre‐pandemic [3] Pre‐pandemic, otherwise nasopharyngeal swab Timing of reference standard: Not stated Blinded to index test: yes, done prior Incorporated index test: No
Flow and timing	Time interval between index and reference tests: [1] Not stated [time of PCR positivity was 5.3 days after date of symptom onset on average (range, 0 to 19 days)]. [2] Not stated [3] The time from an RPP‐positive result to serum collection ranged from 11 to 135 days (mean 45 days) from the date of the original RPP result. All patients received same reference standard: No Missing data: yes (see Tables 3 and 4) Uninterpretable results: Two invalid samples observed for Affinity and for Euroimmun and one for Getein BioTech LFA (all excluded) Indeterminate results: Yes; number of equivocal results reported per test; these can be considered either as index‐positive or negative Unit of analysis: Patients; 11 COVID‐19‐positive patients had serum collected at multiple time periods; however, only one sample per patient was used per time interval to calculate assay sensitivity. When more than one serum sample from the same individual was within a given time interval, only the most recently collected serum sample was included.
Comparative
Notes	Funding: We also thank the following manufacturers for supplying kits for analysis: Abbott, Affinity, Bio‐Rad, DiaSorin, Euroimmun, Roche, BTNX, Biolidics, Deep Blue, Genrui, Getein BioTech, and Innovita. Publication status: Published paper Source: Journal of Clinical Microbiology Author COI: Not stated
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?			Unclear
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?	No
Did all participants receive a reference standard?	Yes
Were results presented per patient?	Yes
Could the patient flow have introduced bias?		High risk