TY - JOUR
T1 - Comparison of in situ hybridization, immunohistochemistry, and reverse transcription-droplet digital polymerase chain reaction for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) testing in tissue
AU - Roden, Anja C.
AU - Vrana, Julie A.
AU - Koepplin, Justin W.
AU - Hudson, Angela E.
AU - Norgan, Andrew P.
AU - Jenkinson, Garrett
AU - Yamaoka, Satoko
AU - Ebihara, Hideki
AU - Monroe, Robert
AU - Szabolcs, Matthias J.
AU - Majumdar, Ramanath
AU - Moyer, Ann M.
AU - García, Joaquín J.
AU - Kipp, Benjamin R.
N1 - Publisher Copyright:
© 2021 College of American Pathologists. All rights reserved.
PY - 2021/7
Y1 - 2021/7
N2 - Context.-Small case series have evaluated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection in formalin-fixed, paraffin-embedded tissue using reverse transcription-polymerase chain reaction, immunohistochemistry (IHC), and/or RNA in situ hybridization (RNAish). Objective.-To compare droplet digital polymerase chain reaction, IHC, and RNAish to detect SARS-CoV-2 in formalin-fixed, paraffin-embedded tissue in a large series of lung specimens from coronavirus disease 2019 (COVID-19) patients. Design.-Droplet digital polymerase chain reaction and RNAish used commercially available probes; IHC used clone 1A9. Twenty-six autopsies of COVID-19 patients with formalin-fixed, paraffin-embedded tissue blocks of 62 lung specimens, 22 heart specimens, 2 brain specimens, and 1 liver, and 1 umbilical cord were included. Control cases included 9 autopsy lungs from patients with other infections/inflammation and virus-infected tissue or cell lines. Results.-Droplet digital polymerase chain reaction had the highest sensitivity for SARS-CoV-2 (96%) when compared with IHC (31%) and RNAish (36%). All 3 tests had a specificity of 100%. Agreement between droplet digital polymerase chain reaction and IHC or RNAish was fair (j ¼ 0.23 and j ¼ 0.35, respectively). Agreement between IHC and in situ hybridization was substantial (j ¼ 0.75). Interobserver reliability was almost perfect for IHC (j ¼ 0.91) and fair to moderate for RNAish (j ¼ 0.38-0.59). Lung tissues from patients who died earlier after onset of symptoms revealed higher copy numbers by droplet digital polymerase chain reaction (P ¼ .03, Pearson correlation ¼-0.65) and were more likely to be positive by RNAish (P ¼ .02) than lungs from patients who died later. We identified SARS-CoV-2 in hyaline membranes, in pneumocytes, and rarely in respiratory epithelium. Droplet digital polymerase chain reaction showed low copy numbers in 7 autopsy hearts from ProteoGenex Inc. All other extrapulmonary tissues were negative. Conclusions.-Droplet digital polymerase chain reaction was the most sensitive and highly specific test to identify SARS-CoV-2 in lung specimens from COVID-19 patients.
AB - Context.-Small case series have evaluated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection in formalin-fixed, paraffin-embedded tissue using reverse transcription-polymerase chain reaction, immunohistochemistry (IHC), and/or RNA in situ hybridization (RNAish). Objective.-To compare droplet digital polymerase chain reaction, IHC, and RNAish to detect SARS-CoV-2 in formalin-fixed, paraffin-embedded tissue in a large series of lung specimens from coronavirus disease 2019 (COVID-19) patients. Design.-Droplet digital polymerase chain reaction and RNAish used commercially available probes; IHC used clone 1A9. Twenty-six autopsies of COVID-19 patients with formalin-fixed, paraffin-embedded tissue blocks of 62 lung specimens, 22 heart specimens, 2 brain specimens, and 1 liver, and 1 umbilical cord were included. Control cases included 9 autopsy lungs from patients with other infections/inflammation and virus-infected tissue or cell lines. Results.-Droplet digital polymerase chain reaction had the highest sensitivity for SARS-CoV-2 (96%) when compared with IHC (31%) and RNAish (36%). All 3 tests had a specificity of 100%. Agreement between droplet digital polymerase chain reaction and IHC or RNAish was fair (j ¼ 0.23 and j ¼ 0.35, respectively). Agreement between IHC and in situ hybridization was substantial (j ¼ 0.75). Interobserver reliability was almost perfect for IHC (j ¼ 0.91) and fair to moderate for RNAish (j ¼ 0.38-0.59). Lung tissues from patients who died earlier after onset of symptoms revealed higher copy numbers by droplet digital polymerase chain reaction (P ¼ .03, Pearson correlation ¼-0.65) and were more likely to be positive by RNAish (P ¼ .02) than lungs from patients who died later. We identified SARS-CoV-2 in hyaline membranes, in pneumocytes, and rarely in respiratory epithelium. Droplet digital polymerase chain reaction showed low copy numbers in 7 autopsy hearts from ProteoGenex Inc. All other extrapulmonary tissues were negative. Conclusions.-Droplet digital polymerase chain reaction was the most sensitive and highly specific test to identify SARS-CoV-2 in lung specimens from COVID-19 patients.
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U2 - 10.5858/arpa.2021-0008-SA
DO - 10.5858/arpa.2021-0008-SA
M3 - Article
C2 - 33720333
AN - SCOPUS:85107577519
SN - 0003-9985
VL - 145
SP - 785
EP - 796
JO - Archives of Pathology and Laboratory Medicine
JF - Archives of Pathology and Laboratory Medicine
IS - 7
ER -