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Enhancing protein signal detection in asexual and viviparous pea aphids: A guided protocol for tissue dissection and proteinase K treatment. | LitMetric

Enhancing protein signal detection in asexual and viviparous pea aphids: A guided protocol for tissue dissection and proteinase K treatment.

MethodsX

Laboratory for Genomics and Development, Department of Entomology, College of Bio-Resources and Agriculture, National Taiwan University (NTU), Taipei, Taiwan.

Published: December 2024

AI Article Synopsis

  • - Aphids reproduce quickly and asexually, and understanding the molecular processes involved requires precise protein detection methods, like immunostaining.
  • - The research aims to optimize proteinase K (PK) digestion to enhance antibody access to various stages of pea aphid embryos and salivary glands.
  • - Detailed protocols for tissue dissection and PK treatment are proposed to improve the accuracy of protein signal detection in aphids.

Article Abstract

Aphids, as hemipteran insects, reproduce via parthenogenesis and viviparity, resulting in rapid and exponential offspring production. To investigate the molecular mechanisms underlying parthenogenetic viviparity in asexual aphids, precise protein detection through immunostaining is essential. Our previous research demonstrated the need for proteinase K (PK) treatment to improve tissue permeability, enabling antibodies targeting the germ-cell marker Ap-Vas1 to access gastrulating and later-stage embryos. However, optimal PK digestion protocols have not been thoroughly explored. In this study, we propose strategies to optimize PK digestion conditions for early, middle, and late-stage pea aphid embryos, which have varying tissue thicknesses. Additionally, we extend the application of PK treatment to salivary glands, a representative somatic tissue, by optimizing conditions for antibody penetration against the salivary gland marker C002. To enhance spatial precision in signal detection, we provide a detailed protocol for tissue dissection specific to pea aphids, focusing on the preservation of tissue integrity. These comprehensive guidelines, covering tissue dissection and PK titration, are expected to improve the specificity and intensity of protein signals in pea aphids and other aphid species.•Provide aphid-specific dissection methods to obtain intact embryos and salivary glands.•Present strategies for optimizing PK treatment conditions across different tissue types.

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Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC11489042PMC
http://dx.doi.org/10.1016/j.mex.2024.102982DOI Listing

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