Acquiring genome‐wide gene expression profiles in Guthrie card blood spots using microarrays

Abstract

Standard Guthrie cards have been widely used to collect blood samples from essentially all USA and Japanese neonates for newborn screening programs. Thus, archival blood spot samples are a unique and comprehensive resource for molecular pathology studies. However, the challenge in using these samples is the presumed low quantity and degraded quality of nucleic acids that can be isolated from these samples, particularly the RNA. Here, we report a new assay using Agilent 4x44K microarrays for acquiring genome-wide gene expression profiles from blood spots on Guthrie cards. Due to the small amount of RNA obtained from each sample, major modifications, such as concentrating and amplifying the RNA and using a different labeling procedure, were performed. Approximately 9000 expressed genes can be detected after normalization of data, an increment of 260% in detection power compared with previously reported cDNA microarrays made in-house with standard procedures. The correlation coefficients in technical and biological replicates were 0.92 and 0.85, respectively, confirming the reproducibility of this study. This new and comprehensive assay will add value to the utility of archival Guthrie cards (e.g. neonatal blood spot cards) and open new opportunities to molecular epidemiology, pathology, genomic, and diagnostic studies of perinatal diseases.

Gene expression profiling of archived dried blood spot samples from the Danish Neonatal Screening Biobank

“…For similar samples tested on the Agilent 44 k array, Khoo et al found a correlation for experimental replicates of R 2 = 0.92 [23]. Overall, the values reported here, R 2 = 0.97 ± 0.01 and R 2 = 0.91 ± 0.02 for different sample groups, are above those of experimental replicates reported as satisfactory for FFPE samples (R 2 = 0.9) [29].…”

“…Other groups have published RNA work using DBS cards as a source [22][23][24]. A series of work by Dr. Khoo and colleagues has convincingly shown that DBS samples from the Michigan Screening Biobank (MSB) can be used for GEP [23] and to detect the differential expression of gender specific genes [25]. Dr. Khoo and her colleagues have also documented differences between DBS stored at room temperature versus conventionally preserved blood after more than 10 years in storage [26].…”

“…Using these approaches, the DNSB has been successfully used to investigate several diseases [16][17][18][19][20][21]. Other groups have published RNA work using DBS cards as a source [22][23][24]. A series of work by Dr. Khoo and colleagues has convincingly shown that DBS samples from the Michigan Screening Biobank (MSB) can be used for GEP [23] and to detect the differential expression of gender specific genes [25].…”

Public concerns regarding the storage and secondary uses of residual newborn bloodspots: an analysis of print media, legal cases, and public engagement activities

“…More recently, genetic technology has "skyrocketed" the research value of RBS (Tarini 2011). For example, researchers successfully amplified DNA and conducted genome-wide gene expression profiles using RBS (Hollegaard et al 2013;Khoo et al 2011). Researchers have also successfully extracted and amplified RNA from stored unfrozen samples almost a decade old (Haak et al 2009).…”

Current status of newborn screening worldwide: 2015

“…With technology moving towards less expensive high-throughput genomic testing, the possibilities of genomic NBS are being discussed, and the ethical, legal, and social implications debated. [66][67][68] In both Canada and the U.S., there is increasing emphasis on the speed with which NBS specimens reach the testing laboratory and have testing results available.…”