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Quantification of rat kisspeptin using a novel radioimmunoassay. | LitMetric

Quantification of rat kisspeptin using a novel radioimmunoassay.

PLoS One

Section of Investigative Medicine, Department of Medicine, Imperial College London, London, United Kingdom.

Published: February 2015

AI Article Synopsis

  • Kisspeptin is a crucial hormone in the hypothalamus that influences puberty and reproductive health, with previous studies using methods like mRNA expression and immunohistochemistry to study it.
  • Despite examining mRNA levels, there’s a gap in understanding how these levels translate to actual protein amounts, leading to challenges in quantifying kisspeptin's presence.
  • A new radioimmunoassay (RIA) was developed for accurately measuring kisspeptin in rodent tissues, confirming its levels in specific brain regions and placenta, and providing a reliable tool for future research on kisspeptin.

Article Abstract

Kisspeptin is a hypothalamic peptide hormone that plays a pivotal role in pubertal onset and reproductive function. Previous studies have examined hypothalamic kisspeptin mRNA expression, either through in situ hybridisation or real-time RT-PCR, as a means quantifying kisspeptin gene expression. However, mRNA expression levels are not always reflected in levels of the translated protein. Kisspeptin-immunoreactivity (IR) has been extensively examined using immunohistochemistry, enabling detection and localisation of kisspeptin perikaya in the arcuate nucleus (ARC) and anteroventral periventricular nucleus (AVPV). However, quantification of kisspeptin-IR remains challenging. We developed a specific rodent radioimmunoassay assay (RIA) capable of detecting and quantifying kisspeptin-IR in rodent tissues. The RIA uses kisspeptin-10 as a standard and radioactive tracer, combined with a commercially available antibody raised to the kisspeptin-10 fragment. Adult female wistar rat brain samples were sectioned at 300 µm and the ARC and AVPV punch micro-dissected. Brain punches were homogenised in extraction buffer and assayed with rodent kisspeptin-RIA. In accord with the pattern of kisspeptin mRNA expression, kisspeptin-IR was detected in both the ARC (47.1±6.2 fmol/punch, mean±SEM n = 15) and AVPV (7.6±1.3 fmol/punch, mean±SEM n = 15). Kisspeptin-IR was also detectable in rat placenta (1.26±0.15 fmol/mg). Reverse phase high pressure liquid chromatography analysis showed that hypothalamic kisspeptin-IR had the same elution profile as a synthetic rodent kisspeptin standard. A specific rodent kisspeptin-RIA will allow accurate quantification of kisspeptin peptide levels within specific tissues in rodent experimental models.

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Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4028310PMC
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0097611PLOS

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