Molecular and Cellular Probes 22 (2008) 76–82 Application of multiplex bead array assay for Yq microdeletion analysis in infertile males Hye-Jung Yeom a,1 , Young-Sun Her a,1 , Moon-Ju Oh a , Saswati Paul a , Mi-Seon Park a , Jong-Pil Yeoun a , Jin-Wook Jung a , Suman Lee b , Seung Yong Hwang a,Ã a Department of Biochemistry, Hanyang University & GenoCheck Co. Ltd., Ansan, Gyeonggi-do 426-791, Republic of Korea b Functional Genomics Lab, CHA Research Institute, Bundang Campus, College of Medicine, Pochon CHA University, 222 Yatap-Dong, Bundang-Gu, Sungnam-Si, Kyunggi-Do 463-836, Republic of Korea Received 21 March 2007; accepted 19 June 2007 Available online 30 June 2007 Abstract The purpose of this study was to apply the multiplex bead array as a diagnostic tool for male infertility. The multiplex bead array offers a new platform in high-throughput nucleic acid detection. Six loci, including sex-determining regions on the Y (SRY) chromosome as a control and five sequence-tagged sites (STS) in azoospermia-factor regions, were used in this system. Extracted genomic DNA from infertile male blood was used for multiplex polymerase chain reaction (PCR). After multiplex PCR using specific Cy3-labeled primer sets, the PCR product was hybridized with capture probes. A multiplexed PCR-liquid bead was arrayed for simultaneous detection using the Luminex system. This assay system correctly identified the presence or deletion of the Y chromosome. Therefore, this method provides a sensitive and high-throughput method for probing the deletion of the Y chromosome, and offers a completely new approach to male infertility screening. r 2007 Elsevier Ltd. All rights reserved. Keywords: Infertile; Luminex; Multiplex PCR; STS markers; Y chromosome 1. Introduction Based on current infertility studies, 10–15% of couples have fertility problems. Male infertility has been identified in 50% of all infertility cases [1,2]. Both qualitative and quantitative abnormalities in sperm production are re- sponsible in 40% of the infertile population. The origin of reduced testicular sperm function in about 60–70% of cases is unknown [3,4]. Several causes of male infertility are known, including azoospermia, oligozoospermia, astheno- zoospermia, teratozoospermia, immunologic factors, and aspermia [5,6]. The long arm of the human Y chromosome is required for sperm production. Deletion in three different regions can cause severe spermatogenic defects, ranging from non- obstructive azoospermia to oligozoospermia. These regions are referred to as ‘azoospermia factors’ (AZFs). Three distinct non-overlapping regions, designated as AZFa, AZFb, and AZFc, are located in interval 5–6 of the Yq chromosome, and are associated with impaired spermato- genesis in humans [7]. Recent studies have shown that it is possible that microdeletions on the Yq chromosome of a father are transmitted to his sons [8]. Therefore, diagnostic detection of a deletion on the Yq chromosome is needed in infertile males hoping to have a baby prior to any fertility treatments. Screening for microdeletions in AZF regions is routinely performed in major andrology and infertility centers [8,9]. However, there is some limit to the applica- tion of these conventional methods in karyotyping, south- ern-blot, and polymerase chain reaction (PCR) in infertility clinics, mainly due to its low specificity, inaccuracy, and time-consuming procedure [10]. Even though most of infertility clinics currently using multiplex PCR rather than single PCR, more precise methodologies are necessary ARTICLE IN PRESS www.elsevier.com/locate/ymcpr 0890-8508/$ - see front matter r 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.mcp.2007.06.004 Ã Corresponding author. Tel.: +82 31 400 5516; fax: +82 31 502 5518. E-mail address: syhwang@hanyang.ac.kr (S.Y. Hwang). 1 These authors contributed equally to this manuscript.