Flow cytometry isolation and reverse transcriptase-polymerase chain reaction characterization of human round spermatids in infertile patients.
نویسندگان
چکیده
Flow cytometry coupled to cell sorting is proposed as a method to isolate round spermatids from testicular biopsies in obstructive azoospermic patients. The cells were separated on the basis of their size and density only. We obtained homogenous populations of alive round spermatids free of lymphocytes and diploid germ cells. The detection of protamine 1 gene (PRM1) and PRM2 expression in the sorted cells proves that these cells are round spermatids. On the contrary, neither the expression of CD3-delta, which is specific to lymphoid cells, nor that of MAGE1, which has been demonstrated in diploid germ cells, could be observed in the round spermatid population even after using a nested polymerase chain reaction (PCR) assay. The flow cytometry procedure failed to isolate round spermatids from ejaculates in non-obstructive azoospermic patients. In > 39 ejaculates tested by reverse transcriptase-PCR, only nine revealed the presence of some round spermatids, as demonstrated by the expression of PRM1. However, these round spermatids did not express PRM2.
منابع مشابه
Flow cytometric method to isolate round spermatids from mouse testis.
The purpose of this study was to isolate pure populations of round spermatids from mouse testis by flow cytometry followed by cell sorting. Cell suspensions from mouse testis were enriched in germ cells by centrifugation on a discontinuous Percoll gradient, then analysed using a FACScalibur flow cytometer measuring the cell size and density. A large and well-delimited population of cells (R1) e...
متن کاملMolecular analysis of the S1 gene of vaccine strains of infectious bronchitis virus using reverse transcriptase-polymerase chain reaction and restriction fragment length polymorphism
Infectious bronchitis virus (IBV) is an acute and contagious viral disease of poultry that affects different systems, including the respiratory tract in particular. IBV causes major economic losses in the poultry industry globally. Due to antigenic variation of the causative agent, control of the disease is difficult. To control the disease, many vaccines that belong to different serotypes are...
متن کاملUnderperformance of Reverse-Transcriptase Polymerase Chain Reaction in Japan and Potential Implications From Diamond Princess Cruise Ship and Other Countries During the Ongoing COVID-19 Pandemic
متن کامل
Detection of Ureaplasma Urealyticum in Clinical Samples from Infertile Women by Polymerase Chain Reaction
Genital Ureaplasma urealyticum infection is considered to be a sexually transmitted infection. The bacterium has been found to be involved in PID, chorioamnionitis, urethritis, respiratory distress syndrome and pneumonia in newborn, abortion and infertility. U. urealyticum infections not only jeopardize fertility but also pose a risk for infertility treatment and resulting pregnancies. The purp...
متن کاملMeasurement by quantitative PCR of changes in HPRT, PGK-1, PGK-2, APRT, MTase, and Zfy gene transcripts during mouse spermatogenesis.
A reverse transcriptase-polymerase chain reaction assay (RT-PCR) was used quantitatively to measure accumulated levels of RNA transcripts in total mouse RNAs derived from male germ cells at various spermatogenic stages. RNA levels for two X-linked enzymes, phosphoglycerate kinase (PGK-1) and hypoxanthine phosphoribosyl transferase (HPRT), both decrease during spermatogenesis, although the trans...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید
ثبت ناماگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید
ورودعنوان ژورنال:
- Human reproduction
دوره 14 2 شماره
صفحات -
تاریخ انتشار 1999