Name: Xenopus Dissection and Perfusion
Uploaded: 2023-05-16T12:50:00
Description: Watch this Scientific Journal Video about Xenopus Dissection and Perfusion at JoVE.com

The video provides a detailed guide on the dissection and perfusion process of a frog (Xenopus), outlining steps such as euthanasia, incision techniques, and heart perfusion to visualize blood flow. It emphasizes monitoring tissue responses and ensuring successful perfusion while noting the visual effects on various organs, especially in albino specimens.

xenopus dissection and perfusion

Blood Perfusion in African Clawed Frogs for Transcriptomics and Proteomics

The whole body perfusion of amphibians is routinely completed for the purposes of preservation and fixation1,2,3,4,5,6. However, these procedures occur at a rate that limits the number of fresh samples that can be taken per animal. The goal of this work is to develop an effective blood perfusion protocol in Xenopus, prioritizing the speed of the technique. The protocol takes less than 10 min per animal for X. tropicalis and less than 15 min per X. laevis animal. The secondary priorities are the ease of replication and the use of easily acquired equipment so that high-quality samples can be shared widely between Xenopus labs.
Xenopus frogs are used widely in biomedical research to study fundamental biological and pathological processes conserved across species. This tetrapod has a closer evolutionary relationship with mammals than other aquatic models, having lungs, a three-chambered heart, and limbs with digits. The international community effectively uses Xenopus to gain a deeper understanding of human disease through in-depth disease modeling and molecular analysis of disease-related gene function. The numerous advantages of Xenopus as an animal model make them invaluable tools to study the molecular basis of human development and disease; these advantages include: large oocyte and embryo size, high fecundity, ease of housing, rapid external development, and ease of genomic manipulation. Xenopus have been estimated to share ~80% of the identified human disease genes7.
Compared to popular mammalian models, Xenopus is a rapid, cost-effective model, with the ease of morpholino knock-down and availability of efficient transgenics and targeted gene mutations using CRISPR8. Quantitative mass spectrometry and single-cell transcriptomics have been successfully applied to Xenopus embryos9,10,&#160;but a recent cell atlas from Xenopus laevis shows that the composition of most tissues is dominated by blood cell types11. By developing a technique that exsanguinates tissue at a rapid rate and using chilled media, the sample freshness is minimally affected by perfusion. This is particularly important for applications where the goal is to profile physiologically unperturbed mRNA or protein expression.

Author Spotlight: Effective Rapid Blood Perfusion in Xenopus  

Effektiv snabb blodperfusion i Xenopus

This procedure centers around the humane utilization of amphibian models in a manner that maximizes efficiency. Standardization practices can be applied to samples taken and preserved at the originating colony, mitigating biosecurity risks and eliminating animal transport stress. Profusion can be completed quickly, maximizing the time available to sample tissues while they are still fresh.This freshness is especially important for proteomic and transcriptomic profiling. This protocol is detailed and explicit, making it usable for individuals with a wide variety of backgrounds. This facilitates the sharing of high-quality samples between Xenopus labs.Along with this profusion technique, a similarly comprehensive protocol is being worked on for the standardized sampling of all major organs.

Watch this Scientific Journal Video about Xenopus Dissection and Perfusion at JoVE.com

Xenopus Dissection and Perfusion  

Xenopus Dissektion och perfusion

Efter avslutad primär eutanasi, kontrollera förlusten av smärtrespons genom att klämma foten. Placera grodan på ryggen. Fäst ner varje lem.Skär genom hudens mittlinje och skär sedan i sidled för att skapa två hudflikar. Ta tag i linea alba med pincett och dra bort den från koelomhålan. Skär genom muskulaturen.Klipp eller fäst alla flikar ur vägen. Skär igenom coracoidbenen med hjälp av dissektionssax och ta bort överflödig vävnad för att få bättre tillgång till hjärtat. Därefter skiftar du försiktigt magen på den dissekerade Xenopus till toppen av vänster leverlob.Ta försiktigt tag i en lungspets med hjälp av vävnadstång och dra den utanför koelomhålan och stift sedan lungan genom spetsen. Dra det tunna perikardiet tätt med vävnadspincett. Använd spetsen på en iridectomy sax, perforera försiktigt perikardiet utan att skära någon underliggande vävnad.Skala perikardiet bort från hjärtkamrarna. Ta försiktigt tag i ventrikelns topp med pincett. Utan att perforera ventrikeln, sätt in nålen genom stängningen av pincetten i ventrikelkammaren.Kläm fast vävnadspincetten på plats med hjälp av en nålhållare eller hemostat. Starta pumpens flöde med cirka fem milliliter per minut. Detta kommer att uppsluka hjärtkamrarna och artärstammen.Använd sax för att göra ett nick i höger öron. Fortsätt pumpflödet tills kärlen i magen blancherar. Lance vänster hjärta öron.Öka pumpens flödeshastighet till 10 ml per minut. Skölj koelomhålan och perfusionsmediet med en överföringspipett. Om organen skiftar under sköljning, flytta dem försiktigt tillbaka för att bibehålla synligheten i magen och lungorna.För att hjälpa till att visualisera perfusatet som strömmar ut ur hjärtat kan auriklarna minskas ytterligare. Håll nålen på plats tills magen, kärlen och lungorna har blancherat och perfusatet som strömmar från hjärtat verkar klart. Framgångsrik perfusion observerades i alla vävnader utom levern, vilket gjorde dem tydligt lättare och mindre mättade med blod.Detta är mest synligt i albinos.

Research

JoVE Journal

Biology

Xenopus Dissection and Perfusion

Effective Rapid Blood Perfusion in Xenopus

Xenopus have been powerful model organisms for understanding vertebrate development and disease for over 100 years. Here, a rapid blood perfusion protocol ...