PhD Scientific Days 2024

Poster Session A - Molecular Medicine 1.

Optimization of Sirius Red-Based Microplate Assay to Investigate Collagen Production In Vitro

Előadó neve

Veres-Székely, Apor, PhD

Neptun code

KYS71E

Előadó munkahelye

Pediatric Center, Semmelweis University; HUN-REN-SU Pediatrics and Nephrology Research Group

Előadó telefonszáma

+36 1 459 1500/52710

Előadó e-mail címe

veres-szekely.apor@semmelweis.hu

Az előadás címe

Optimization of Sirius Red-Based Microplate Assay to Investigate Collagen Production In Vitro

Szerző(k) neve és munkahelye

Apor Veres-Székely¹, Csenge Szász², Domonkos Pap¹, Beáta Szebeni¹, Péter Bokrossy², László Őrfi³, Attila J. Szabó¹, Ádám Vannay¹

1: Pediatric Center, Semmelweis University; HUN-REN-SU Pediatrics and Nephrology Research Group
2: Pediatric Center, Semmelweis University
3: Department of Pharmaceutical Chemistry, Semmelweis University; Vichem Chemie Ltd.

Bemutatás módja

Poszter

Szekció

Poster Session A - Molecular Medicine 1.

Language of the presentation

Hungarian

Preferred session

Molecular Medicine

Összefoglaló szövege

Introduction: Tissue fibrosis is characterized by chronic fibroblast activation and consequently excessive accumulation of collagen-rich extracellular matrix. In vitro microplate-based assays are essential to investigate the underlying mechanism and the effect of antifibrotic drugs.
Method: In this study, in the absence of a gold-standard method, we optimized a simple, cost-effective, Sirius Red-based colorimetric measurement to determine the collagen production of fibroblasts grown on 96-well tissue culture plates.
Results: Based on our findings, the use of a serum-free medium is recommended to avoid aspecific signals, while ascorbate supplementation increases the collagen production of fibroblasts. The cell-associated collagens can be quantified by Sirius Red staining in acidic conditions followed by alkaline elution. Immature collagens can be precipitated from the culture medium by acidic Sirius Red solution, and after subsequent centrifugation and washing steps, their amount can be also measured. Increased attention has been paid to optimizing the assay procedure, including incubation time, temperature, and solution concentrations.
Conclusion: The resulting assay shows high linearity and sensitivity and could serve as a useful tool in fibrosis-related basic research as well as in preclinical drug screening.
Funding: This research was founded by the National Research, Development and Innovation Office (NKFIH), K-142728; Semmelweis University, TKP2021-EGA-24, STIA-KFI-2021; Hungarian Research Network, ELKH-POC-2022-024, the New National Excellence Program of the Ministry for Culture and Innovation from the Source of the National Research, Development and Innovation Fund, ÚNKP-23-3-I-SE-36, ÚNKP-23-3-I-SE-42, ÚNKP-23-4-II-SE-29, ÚNKP-23-5-SE-15; Hungarian Academy of Sciences, János Bolyai Research Scholarship.

University

Semmelweis University

Supervisor

-

Publication of my abstract

I do not give consent to the publication of my abstract on the website of the congress.

Kind

Szabad

Status

elfogadva

Accepted presentation method

poszter

Előadás fájl jóváhagyás

nem rendelkezett róla

Előadó

2965

Start

15:00

End

15:03