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Comparison of two techniques for assessing pancreatic islet viability: flow cytometry and fluorescein diacetate/propidium iodine staining
Lemos, Natália Emerim; Dieter, Cristine; Farias, Mariela Granero; Rheinheimer, Jakeline; Souza, Bianca Marmontel de; Bauer, Andrea Carla; Crispim, Daisy.
  • Lemos, Natália Emerim; Hospital de Clínicas de Porto Alegre. Serviço de Endocrinologia. Laboratório de Biologia das Ilhotas Pancreáticas Humanas. Porto Alegre. BR
  • Dieter, Cristine; Hospital de Clínicas de Porto Alegre. Serviço de Endocrinologia. Laboratório de Biologia das Ilhotas Pancreáticas Humanas. Porto Alegre. BR
  • Farias, Mariela Granero; Hospital de Clínicas de Porto Alegre. Serviço de Diagnóstico Laboratorial. Unidade de Diagnóstico Especializado. Porto Alegre. BR
  • Rheinheimer, Jakeline; Hospital de Clínicas de Porto Alegre. Serviço de Endocrinologia. Laboratório de Biologia das Ilhotas Pancreáticas Humanas. Porto Alegre. BR
  • Souza, Bianca Marmontel de; Hospital de Clínicas de Porto Alegre. Serviço de Endocrinologia. Laboratório de Biologia das Ilhotas Pancreáticas Humanas. Porto Alegre. BR
  • Bauer, Andrea Carla; Hospital de Clínicas de Porto Alegre. Serviço de Endocrinologia. Laboratório de Biologia das Ilhotas Pancreáticas Humanas. Porto Alegre. BR
  • Crispim, Daisy; Hospital de Clínicas de Porto Alegre. Serviço de Endocrinologia. Laboratório de Biologia das Ilhotas Pancreáticas Humanas. Porto Alegre. BR
Clin. biomed. res ; 41(4): 325-331, 2021. ilus
Article in English | LILACS | ID: biblio-1349503
ABSTRACT

Introduction:

The success of islet transplantation for patients with unstable type 1 diabetes mellitus depends, in part, on the number of isolated islets and their quality, which is assessed by functional and viability tests. The test currently employed to evaluate islet viability, used by the Collaborative Islet Transplant Registry to release products for transplantation, is fluorescein diacetate/propidium iodide (FDA/PI) staining. However, the efficacy of this method relies on researcher experience; in this context, a quantitative method may be useful. The aim of this study was to compare islet viability as assessed by flow cytometry and the FDA/PI assay.

Methods:

Viability was analyzed in islets isolated from 10 male Wistar rats. Upon FDA/PI staining, 50 islets from each animal were analyzed under fluorescence microscopy by two well-trained researchers. For flow cytometry, islets were dispersed and 100 000 single cells were incubated with the 7-amino-actinomycin D (7AAD) fluorophore (dyes necrotic and late apoptotic cells) and the Annexin V-APC antibody (marks early apoptotic cells).

Results:

A moderate correlation was found between techniques (r = 0.6; p = 0.047). The mean islet viability measured by flow cytometry was higher than that estimated using FDA/PI staining (95.5 ± 1.4% vs 89.5 ± 5.0%; p = 0.002).

Conclusions:

Although flow cytometry is more expensive and time-consuming than FDA/PI staining, it is a quantitative technique with greater reproducibility that is less subject to inter-observer variability than FDA/PI. Therefore, flow cytometry appears to be the technique of choice when aiming for a more precise determination of islet viability. (AU)
Subject(s)


Full text: Available Index: LILACS (Americas) Main subject: Propidium / Islets of Langerhans Transplantation / Fluorescein / Flow Cytometry Limits: Animals Language: English Journal: Clin. biomed. res Journal subject: Medicine Year: 2021 Type: Article Affiliation country: Brazil Institution/Affiliation country: Hospital de Clínicas de Porto Alegre/BR

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Full text: Available Index: LILACS (Americas) Main subject: Propidium / Islets of Langerhans Transplantation / Fluorescein / Flow Cytometry Limits: Animals Language: English Journal: Clin. biomed. res Journal subject: Medicine Year: 2021 Type: Article Affiliation country: Brazil Institution/Affiliation country: Hospital de Clínicas de Porto Alegre/BR