Pesenti 2017.

Study characteristics
Patient Sampling	Inclusion/exclusion criteria Inclusion criteria: diffuse glioma; availability of tumour and peripheral blood specimens Prior testing Presumably histopathological diagnosis
Patient characteristics and setting	Number of participants/tumours with results for 1p/19q status by ≥ 2 DNA‐based tests: 50 Country: Italy Population source and setting: Fondazione IRCCS Ca' Granda, Ospedale Maggiore Policlinico di Milano, Italy. December 2013 to November 2016 Agea: median: 53 years, interquartile range: NR; range: 21–81 years Gender: 56.0% male Karnofsky performance status: NR First diagnosis/recurrent disease: NR aFor whole population: all 50 participants had 1 tests (MS and PCR‐based LOH), only a subset had aCGH or FISH.
Index tests	4 tests: aCGH, FISH, MS and PCR‐based LOH aCGH Tumour sample type: FFPE Region(s) analysed: genome wide Cut‐off: quote: "The aberration filter was set to detect a minimum of five consecutive probes/region, and the minimum absolute average log ratio (MAALR) was ± 0.25. A second analysis was run with a MAALR of ± 0.15 (again with a minimum number of five probes/region), to detect low level mosaicism". Additional details: SurePrint G3 Human CGH 4 × 180K, Agilent Technologies, Santa Clara, California, USA MS Tumour sample type: FFPE Region(s) analysed: rs3737577 (1p21.2), rs59317557 (1p21.2), rs2038366 (1p21.2), rs859104 (1p21.3), rs17378384 (1p31.3), rs2455638 (1p32.1), rs550663 (1p33), rs586057 (1p34.3), rs624971 (1p34.3), rs16866144 (1p35.2), rs11247639 (1p36.11), rs2473287 (1p36.12), rs7512426 (1p36.21), rs809972 (1p36.22), rs4908744 (1p36.23), rs6426368 (1p36.32), rs28503746 (19q13.2), rs67421541 (19q13.2), rs12611404 (19q13.2), rs6070 (19q13.33), rs1674139 (19q13.33), rs1807277 (19q13.33), rs186585 (19q13.41), rs28702875 (19q13.41), rs11666952 (19q13.42), rs36629 (19q13.42) and rs437229 (19q13.43) Cut‐off: the following equation was used to quantitatively define the LOH status: (N2/N1)/((N2/N1) + (T2/T1)) where N1 and N2 were the frequencies of Allele 1 and Allele 2 found in peripheral blood lymphocyte DNA and T1 and T2 were those of the corresponding alleles in tumour DNA. LOH was defined as detected with the value obtained using this formula was < 0.3 or > 0.7. LOH/NO LOH status was defined by the presence of ≥ 2 informative SNPs per chromosome arm with concordant results, 1 of which was located in a centromeric region and the other at a telomeric locus. Additional details: MassARRAY iPLEX platform (Agena Bioscience, San Diego, California, USA), based on MALDI‐TOF MS. PCR performed as a first step. PCR Tumour sample type: FFPE Region(s) analysed: D1S1592 (1p36.13), D1S548 (1p36.23), D1S2694 (1p36.23), D1S2666 (1p36.23), D1S1612 (1p36.23), D1S468 (1p36.32), D19S412 (19q13.32), D19S596 (19q13.33) and D19S206 (19q13.41) Cut‐off: the peak height derived from each allele amplified from both tumor and corresponding normal DNA was compared. The formula (T1/T2)/(N1/N2) was applied, where T1 and T2 were the peak heights of the alleles detected in tumor DNA, and N1 and N2 were the peak heights produced from peripheral blood lymphocyte DNA. LOH was considered present when the result of the calculation was < 0.50. For values > 1.00, the ratio was converted to 1/[(T1/T2)/(N1/N2)] and, again, LOH was considered present if the resulting value was < 0.50. Additional details: analysed by capillary gel electrophoresis using Gene Mapper software on an ABI 3130XL system FISH Tumour sample type: FFPE Region(s) analysed: p36/1q25 and 19q13/19p13 (ZytoVision, Bremerhaven, Germany) Cut‐off: quote: "Interpretation of FISH images was performed accordingly to Ambros et al, 2001 [37]: normal pattern was defined by the presence of an equal number of control/green and target/red signals (i.e. control/target ratio: 2/2, 3/3, 4/4, etc), deletion pattern was characterized by the presence of at least two control/green signals but only one or zero target/red signals (i.e. control/target ratio: 2/1, 2/0, 3/1, etc); finally imbalance pattern was identified by the presence of more than 1 target/red signal (i.e. control/target ratio: 3/2, 4/2, 4/3, etc). A sample was considered positive for 1p/19q codeletion when more than 50% of nuclei per chromosome arm displayed a typical deletion pattern".
Target condition and reference standard(s)	Target condition was absolute 1p/19q deletion. FISH or PCR‐based LOH used as reference standard in some of our analyses.
Flow and timing	We presumed that all tests were performed on biopsied tumour material collected on 1 occasion.
Comparative
Notes
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?	Yes
Did the study avoid inappropriate exclusions?	Unclear
Could the selection of patients have introduced bias?		Unclear risk
Are there concerns that the included patients and setting do not match the review question?			Low concern
DOMAIN 2: Index Test (NanoString)
DOMAIN 2: Index Test (aCGH)
If a threshold was used, was it pre‐specified?	Yes
Were the index test results interpreted without knowledge of the results of the other tests being compared?	Yes
Could the conduct or interpretation of the index test have introduced bias?		Low risk
Are there concerns that the index test, its conduct, or interpretation differ from the review question?			Low concern
DOMAIN 2: Index Test (NGS)
DOMAIN 2: Index Test (G‐banding)
DOMAIN 2: Index Test (FISH (variant 4))
DOMAIN 2: Index Test (SNP array)
DOMAIN 2: Index Test (PCR (with comparison to normal DNA))
DOMAIN 2: Index Test (PCR (without comparison to normal DNA))
DOMAIN 2: Index Test (CISH)
DOMAIN 2: Index Test (MS)
If a threshold was used, was it pre‐specified?	Yes
Were the index test results interpreted without knowledge of the results of the other tests being compared?	Yes
Could the conduct or interpretation of the index test have introduced bias?		Low risk
Are there concerns that the index test, its conduct, or interpretation differ from the review question?			Low concern
DOMAIN 2: Index Test (RFLP)
DOMAIN 2: Index Test (PCR‐based LOH)
If a threshold was used, was it pre‐specified?	Unclear
Were the index test results interpreted without knowledge of the results of the other tests being compared?	Unclear
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 2: Index Test (NGS or aCGH (or both))
DOMAIN 2: Index Test (Methylation array)
DOMAIN 2: Index Test (FISH)
If a threshold was used, was it pre‐specified?	Yes
Were the index test results interpreted without knowledge of the results of the other tests being compared?	Unclear
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 2: Index Test (FISH (variant 1))
DOMAIN 2: Index Test (FISH (variant 2))
DOMAIN 2: Index Test (FISH (variant 3))
DOMAIN 2: Index Test (Real‐time PCR)
DOMAIN 2: Index Test (MLPA)
DOMAIN 2: Index Test (CGH)
DOMAIN 3: Reference Standard
Is the reference standards likely to correctly classify the target condition?	No
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?			Low concern
DOMAIN 4: Flow and Timing
Was there an appropriate interval between index test and reference standard?	Yes
Were all patients included in the analysis?	No
Could the patient flow have introduced bias?		High risk