Dataset Information

Histological cut of a paraffin-embedded blastocyst: Optimized protocol for murine blastocysts.

ABSTRACT: Paraffin-embedded tissues have been used for research and therapeutic applications for decades, as they represent a valuable tool in histology and for molecular analysis, as well as being a way to preserve tissue samples for long periods at a low cost. For tissues such as the liver, lungs, kidney, heart or brain, there are many protocols available, already optimized. The purpose of this work is to optimize and simplify the protocols already available to take a single blastocyst from a mouse, fix it and embed it into a paraffin block without using gelatin, to then perform histological cuts using a microtome, with no need of sophisticated equipment or trained personnel. •The protocol presented here preserves well the morphology of the blastocyst.•Paraffin-embedded sections of the sample can be used for studies such as in situ hybridization, immunohistochemistry, enzyme histochemistry, DNA, RNA or protein extractions, analysis of biomarkers, characterization of surface markers of stem cells integrated into the embryo, to prepare histological material for educational purposes, etc.•Some of these studies could represent a valuable source of new information for the field of reproductive biology.

SUBMITTER: Gracia AU

PROVIDER: S-EPMC7794613 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Publications

Histological cut of a paraffin-embedded blastocyst: Optimized protocol for murine blastocysts.

Gracia Alejandra Usón AU López-Morató Marta M Mijares José J Sánchez-Mateos Soledad S Sánchez-Margallo Francisco Miguel FM Álvarez Ignacio Santiago IS Hernández Nuria N

MethodsX 20191216

Paraffin-embedded tissues have been used for research and therapeutic applications for decades, as they represent a valuable tool in histology and for molecular analysis, as well as being a way to preserve tissue samples for long periods at a low cost. For tissues such as the liver, lungs, kidney, heart or brain, there are many protocols available, already optimized. The purpose of this work is to optimize and simplify the protocols already available to take a single blastocyst from a mouse, fix ...[more]

PMID: 33457212

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Project description:Ticks are important ectoparasites that are capable of transmitting multiple classes of pathogens and are currently linked with many emerging tick-borne diseases worldwide. With increasing occurrences of tick-borne diseases in both humans and veterinary species, there is a continuous need to further our understanding of ticks and the pathogens they transmit. Whole tick histology provides a full scope of the tick internal anatomy, allowing researchers to examine multiple organs of interest in a single section. This is in contrast to other techniques that are more commonly utilized in tick-borne disease research, such as electron microscopy and light microscopy of individual organs. There is a lack of literature describing a practical technique to process whole tick histologic sections. Therefore, the current study aims to provide researchers with a workable protocol to prepare high quality paraffin-embedded whole tick histology sections. Amblyomma americanum adults were used as an example species for this study. After a series of pilot experiments using a combination of various fixatives, softening agents and processing techniques, we elected to compare two common fixatives, 10% neutral-buffered formalin (NBF) and Bouin's solution for whole ticks. Equal numbers of A. americanum unfed adults (n = 10/fixative) were processed identically and their whole tick histology coronal sections were individually scored. Higher scores were assigned to whole tick sections that contained more internal organs that are crucial for tick-borne disease research (e.g. salivary glands and midgut), high integrity of tissues and exoskeleton on the section, and good fixation and staining quality of the tissues. The mean total scores for Bouin's-fixed ticks were significantly higher compared to NBF-fixed ticks (p = 0.001). To further assess our preferred technique, we also demonstrated the feasibility of producing high quality whole tick sections for three other common tick species of medical importance (Rhipicephalus sanguineus, Ixodes scapularis, and Dermacentor variabilis) using Bouin's solution. While this technique may require further optimization for other tick species, we described a feasible protocol that uses commonly available tools, reagents and standard histologic equipment. This should allow any investigator to easily make adjustments to this protocol as needed based on their experimental goals.

Dataset Information

Histological cut of a paraffin-embedded blastocyst: Optimized protocol for murine blastocysts.

Publications

Histological cut of a paraffin-embedded blastocyst: Optimized protocol for murine blastocysts.

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