Multicenter evaluation of a standardized protocol for noninvasive gene expression profiling

K. S. Keslar, M. Lin, A. A. Zmijewska, T. K. Sigdel, T. Q. Tran, L. Ma, M. Bhasin, P. Rao, R. Ding, D. N. Iklé, R. B. Mannon, M. M. Sarwal, T. B. Strom, E. F. Reed, P. S. Heeger, Manikkam Suthanthiran, R. L. Fairchild

Research output: Contribution to journalArticle

28 Citations (Scopus)

Abstract

Gene expression profiling of transplant recipient blood and urine can potentially be used to monitor graft function, but the multitude of protocols in use make sharing data and comparing results from different laboratories difficult. The goal of this study was to evaluate the performance of current methods of RNA isolation, reverse transcription and quantitative polymerase chain reaction (qPCR) and to test whether multiple centers using a standardized protocol can obtain the same results. Samples, reagents and detailed instructions were distributed to six participating sites that performed RNA isolation, reverse transcription and qPCR for 18S, PRF, GZB, IL8, CXCL9 and CXCL10 as instructed. All data were analyzed at a single site. All sites demonstrated proficiency in RNA isolation and qPCR analysis. Gene expression measurements for all targets and samples had correlations >0.938. The coefficient of variation of fold-changes between pairs of samples was less than 40%. All sites were able to accurately quantify a control sample of known concentration within a factor of 1.5. Collectively, we have formulated and validated detailed methods for measuring gene expression in blood and urine that can yield consistent results in multiple laboratories.

Original languageEnglish
Pages (from-to)1891-1897
Number of pages7
JournalAmerican Journal of Transplantation
Volume13
Issue number7
DOIs
Publication statusPublished - 1 Jul 2013
Externally publishedYes

Fingerprint

Gene Expression Profiling
RNA
Polymerase Chain Reaction
Reverse Transcription
Urine
Gene Expression
Information Dissemination
Interleukin-8
Transplants

Keywords

  • Allograft monitoring
  • gene expression profiling
  • kidney allograft
  • multicenter studies
  • quantitative mRNA
  • quantitative PCR

ASJC Scopus subject areas

  • Immunology and Allergy
  • Transplantation
  • Pharmacology (medical)

Cite this

Keslar, K. S., Lin, M., Zmijewska, A. A., Sigdel, T. K., Tran, T. Q., Ma, L., ... Fairchild, R. L. (2013). Multicenter evaluation of a standardized protocol for noninvasive gene expression profiling. American Journal of Transplantation, 13(7), 1891-1897. https://doi.org/10.1111/ajt.12284

Multicenter evaluation of a standardized protocol for noninvasive gene expression profiling. / Keslar, K. S.; Lin, M.; Zmijewska, A. A.; Sigdel, T. K.; Tran, T. Q.; Ma, L.; Bhasin, M.; Rao, P.; Ding, R.; Iklé, D. N.; Mannon, R. B.; Sarwal, M. M.; Strom, T. B.; Reed, E. F.; Heeger, P. S.; Suthanthiran, Manikkam; Fairchild, R. L.

In: American Journal of Transplantation, Vol. 13, No. 7, 01.07.2013, p. 1891-1897.

Research output: Contribution to journalArticle

Keslar, KS, Lin, M, Zmijewska, AA, Sigdel, TK, Tran, TQ, Ma, L, Bhasin, M, Rao, P, Ding, R, Iklé, DN, Mannon, RB, Sarwal, MM, Strom, TB, Reed, EF, Heeger, PS, Suthanthiran, M & Fairchild, RL 2013, 'Multicenter evaluation of a standardized protocol for noninvasive gene expression profiling', American Journal of Transplantation, vol. 13, no. 7, pp. 1891-1897. https://doi.org/10.1111/ajt.12284
Keslar, K. S. ; Lin, M. ; Zmijewska, A. A. ; Sigdel, T. K. ; Tran, T. Q. ; Ma, L. ; Bhasin, M. ; Rao, P. ; Ding, R. ; Iklé, D. N. ; Mannon, R. B. ; Sarwal, M. M. ; Strom, T. B. ; Reed, E. F. ; Heeger, P. S. ; Suthanthiran, Manikkam ; Fairchild, R. L. / Multicenter evaluation of a standardized protocol for noninvasive gene expression profiling. In: American Journal of Transplantation. 2013 ; Vol. 13, No. 7. pp. 1891-1897.
@article{f6b252beca8549508a1f6c9e8c92b680,
title = "Multicenter evaluation of a standardized protocol for noninvasive gene expression profiling",
abstract = "Gene expression profiling of transplant recipient blood and urine can potentially be used to monitor graft function, but the multitude of protocols in use make sharing data and comparing results from different laboratories difficult. The goal of this study was to evaluate the performance of current methods of RNA isolation, reverse transcription and quantitative polymerase chain reaction (qPCR) and to test whether multiple centers using a standardized protocol can obtain the same results. Samples, reagents and detailed instructions were distributed to six participating sites that performed RNA isolation, reverse transcription and qPCR for 18S, PRF, GZB, IL8, CXCL9 and CXCL10 as instructed. All data were analyzed at a single site. All sites demonstrated proficiency in RNA isolation and qPCR analysis. Gene expression measurements for all targets and samples had correlations >0.938. The coefficient of variation of fold-changes between pairs of samples was less than 40{\%}. All sites were able to accurately quantify a control sample of known concentration within a factor of 1.5. Collectively, we have formulated and validated detailed methods for measuring gene expression in blood and urine that can yield consistent results in multiple laboratories.",
keywords = "Allograft monitoring, gene expression profiling, kidney allograft, multicenter studies, quantitative mRNA, quantitative PCR",
author = "Keslar, {K. S.} and M. Lin and Zmijewska, {A. A.} and Sigdel, {T. K.} and Tran, {T. Q.} and L. Ma and M. Bhasin and P. Rao and R. Ding and Ikl{\'e}, {D. N.} and Mannon, {R. B.} and Sarwal, {M. M.} and Strom, {T. B.} and Reed, {E. F.} and Heeger, {P. S.} and Manikkam Suthanthiran and Fairchild, {R. L.}",
year = "2013",
month = "7",
day = "1",
doi = "10.1111/ajt.12284",
language = "English",
volume = "13",
pages = "1891--1897",
journal = "American Journal of Transplantation",
issn = "1600-6135",
publisher = "Wiley-Blackwell",
number = "7",

}

TY - JOUR

T1 - Multicenter evaluation of a standardized protocol for noninvasive gene expression profiling

AU - Keslar, K. S.

AU - Lin, M.

AU - Zmijewska, A. A.

AU - Sigdel, T. K.

AU - Tran, T. Q.

AU - Ma, L.

AU - Bhasin, M.

AU - Rao, P.

AU - Ding, R.

AU - Iklé, D. N.

AU - Mannon, R. B.

AU - Sarwal, M. M.

AU - Strom, T. B.

AU - Reed, E. F.

AU - Heeger, P. S.

AU - Suthanthiran, Manikkam

AU - Fairchild, R. L.

PY - 2013/7/1

Y1 - 2013/7/1

N2 - Gene expression profiling of transplant recipient blood and urine can potentially be used to monitor graft function, but the multitude of protocols in use make sharing data and comparing results from different laboratories difficult. The goal of this study was to evaluate the performance of current methods of RNA isolation, reverse transcription and quantitative polymerase chain reaction (qPCR) and to test whether multiple centers using a standardized protocol can obtain the same results. Samples, reagents and detailed instructions were distributed to six participating sites that performed RNA isolation, reverse transcription and qPCR for 18S, PRF, GZB, IL8, CXCL9 and CXCL10 as instructed. All data were analyzed at a single site. All sites demonstrated proficiency in RNA isolation and qPCR analysis. Gene expression measurements for all targets and samples had correlations >0.938. The coefficient of variation of fold-changes between pairs of samples was less than 40%. All sites were able to accurately quantify a control sample of known concentration within a factor of 1.5. Collectively, we have formulated and validated detailed methods for measuring gene expression in blood and urine that can yield consistent results in multiple laboratories.

AB - Gene expression profiling of transplant recipient blood and urine can potentially be used to monitor graft function, but the multitude of protocols in use make sharing data and comparing results from different laboratories difficult. The goal of this study was to evaluate the performance of current methods of RNA isolation, reverse transcription and quantitative polymerase chain reaction (qPCR) and to test whether multiple centers using a standardized protocol can obtain the same results. Samples, reagents and detailed instructions were distributed to six participating sites that performed RNA isolation, reverse transcription and qPCR for 18S, PRF, GZB, IL8, CXCL9 and CXCL10 as instructed. All data were analyzed at a single site. All sites demonstrated proficiency in RNA isolation and qPCR analysis. Gene expression measurements for all targets and samples had correlations >0.938. The coefficient of variation of fold-changes between pairs of samples was less than 40%. All sites were able to accurately quantify a control sample of known concentration within a factor of 1.5. Collectively, we have formulated and validated detailed methods for measuring gene expression in blood and urine that can yield consistent results in multiple laboratories.

KW - Allograft monitoring

KW - gene expression profiling

KW - kidney allograft

KW - multicenter studies

KW - quantitative mRNA

KW - quantitative PCR

UR - http://www.scopus.com/inward/record.url?scp=84879540321&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84879540321&partnerID=8YFLogxK

U2 - 10.1111/ajt.12284

DO - 10.1111/ajt.12284

M3 - Article

VL - 13

SP - 1891

EP - 1897

JO - American Journal of Transplantation

JF - American Journal of Transplantation

SN - 1600-6135

IS - 7

ER -