Non-radioactive labelling of calcium oxalate crystals for investigations of crystal-cell interactions and internalization

The study of interactions between renal tubular cells and calcium oxalate (CaOx) crystals and their internalization was limited in the past due to lack of a simple method for visualization of CaOx crystals during such processes. We have developed non-radioactive techniques for efficiently labelling...

Full description

Saved in:
Bibliographic Details
Main Authors: Sakdithep Chaiyarit, Siriwan Mungdee, Visith Thongboonkerd
Other Authors: Mahidol University
Format: Article
Published: 2018
Subjects:
Online Access:https://repository.li.mahidol.ac.th/handle/123456789/28889
Tags: Add Tag
No Tags, Be the first to tag this record!
Institution: Mahidol University
id th-mahidol.28889
record_format dspace
spelling th-mahidol.288892018-09-24T15:59:44Z Non-radioactive labelling of calcium oxalate crystals for investigations of crystal-cell interactions and internalization Sakdithep Chaiyarit Siriwan Mungdee Visith Thongboonkerd Mahidol University Chemical Engineering Chemistry Engineering The study of interactions between renal tubular cells and calcium oxalate (CaOx) crystals and their internalization was limited in the past due to lack of a simple method for visualization of CaOx crystals during such processes. We have developed non-radioactive techniques for efficiently labelling and imaging CaOx crystals in the study of crystal-cell interactions and internalization. A total of 12 ionic dyes, as well as AlexaFluor-488, FITC-conjugated IgG and Cy3-conjugated IgG were used to stain/label CaOx crystals. Thereafter, the crystals were incubated with MDCK cells for 48 h. The crystal images were obtained using light, phase-contrast, fluorescence, or laser-scanning confocal microscopy. The internalized CaOx crystals were finally quantified by flow cytometry. From 12 ionic dyes tested, CaOx monohydrate (COM) crystals were stainable only with CBB-R250 (blue) and Ponceau-S (red), whereas CaOx dihydrate (COD) crystals were stainable only with CBB-R250 (blue) and CBB-G250 (blue), which did not stain COM crystals but transformed them to COD. Additionally, only COM could be labelled and imaged with AlexaFluor-488 (green), FITC-conjugated IgG (green) and Cy3-conjugated IgG (red). Crystal-cell interactions (indicated by interrupted borders of crystals) and adhesion were successfully visualized under a light, phase-contrast, or fluorescence microscope. Moreover, laser-scanning confocal microscopic examination successfully identified internalized crystals, which could be quantified by flow cytometry. These non-radioactive techniques are very simple and effective for labelling and imaging COM and COD crystals for the study of crystal-cell interactions, adhesion and internalization, and will be very useful to investigate mechanisms of kidney stone formation. © 2010 The Royal Society of Chemistry. 2018-09-24T08:51:24Z 2018-09-24T08:51:24Z 2010-10-01 Article Analytical Methods. Vol.2, No.10 (2010), 1536-1541 10.1039/c0ay00321b 17599679 17599660 2-s2.0-78650435940 https://repository.li.mahidol.ac.th/handle/123456789/28889 Mahidol University SCOPUS https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=78650435940&origin=inward
institution Mahidol University
building Mahidol University Library
continent Asia
country Thailand
Thailand
content_provider Mahidol University Library
collection Mahidol University Institutional Repository
topic Chemical Engineering
Chemistry
Engineering
spellingShingle Chemical Engineering
Chemistry
Engineering
Sakdithep Chaiyarit
Siriwan Mungdee
Visith Thongboonkerd
Non-radioactive labelling of calcium oxalate crystals for investigations of crystal-cell interactions and internalization
description The study of interactions between renal tubular cells and calcium oxalate (CaOx) crystals and their internalization was limited in the past due to lack of a simple method for visualization of CaOx crystals during such processes. We have developed non-radioactive techniques for efficiently labelling and imaging CaOx crystals in the study of crystal-cell interactions and internalization. A total of 12 ionic dyes, as well as AlexaFluor-488, FITC-conjugated IgG and Cy3-conjugated IgG were used to stain/label CaOx crystals. Thereafter, the crystals were incubated with MDCK cells for 48 h. The crystal images were obtained using light, phase-contrast, fluorescence, or laser-scanning confocal microscopy. The internalized CaOx crystals were finally quantified by flow cytometry. From 12 ionic dyes tested, CaOx monohydrate (COM) crystals were stainable only with CBB-R250 (blue) and Ponceau-S (red), whereas CaOx dihydrate (COD) crystals were stainable only with CBB-R250 (blue) and CBB-G250 (blue), which did not stain COM crystals but transformed them to COD. Additionally, only COM could be labelled and imaged with AlexaFluor-488 (green), FITC-conjugated IgG (green) and Cy3-conjugated IgG (red). Crystal-cell interactions (indicated by interrupted borders of crystals) and adhesion were successfully visualized under a light, phase-contrast, or fluorescence microscope. Moreover, laser-scanning confocal microscopic examination successfully identified internalized crystals, which could be quantified by flow cytometry. These non-radioactive techniques are very simple and effective for labelling and imaging COM and COD crystals for the study of crystal-cell interactions, adhesion and internalization, and will be very useful to investigate mechanisms of kidney stone formation. © 2010 The Royal Society of Chemistry.
author2 Mahidol University
author_facet Mahidol University
Sakdithep Chaiyarit
Siriwan Mungdee
Visith Thongboonkerd
format Article
author Sakdithep Chaiyarit
Siriwan Mungdee
Visith Thongboonkerd
author_sort Sakdithep Chaiyarit
title Non-radioactive labelling of calcium oxalate crystals for investigations of crystal-cell interactions and internalization
title_short Non-radioactive labelling of calcium oxalate crystals for investigations of crystal-cell interactions and internalization
title_full Non-radioactive labelling of calcium oxalate crystals for investigations of crystal-cell interactions and internalization
title_fullStr Non-radioactive labelling of calcium oxalate crystals for investigations of crystal-cell interactions and internalization
title_full_unstemmed Non-radioactive labelling of calcium oxalate crystals for investigations of crystal-cell interactions and internalization
title_sort non-radioactive labelling of calcium oxalate crystals for investigations of crystal-cell interactions and internalization
publishDate 2018
url https://repository.li.mahidol.ac.th/handle/123456789/28889
_version_ 1763489443544563712