Diagnostic filipin testing

Dita Musalkova; Filip Majer; Ladislav Kuchar; Ondrej Luksan; Befekadu Asfaw; Hana Vlaskova; Gabriela Storkanova; Martin Reboun; Helena Poupetova; Helena Jahnova; Helena Hulkova; Jana Ledvinova; Lenka Dvorakova; Jakub Sikora; Milan Jirsa; Marie T. Vanier; Martin Hrebicek

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Diagnostic filipin testing

DM Dita Musalkova

FM Filip Majer

LK Ladislav Kuchar

OL Ondrej Luksan

BA Befekadu Asfaw

HV Hana Vlaskova

GS Gabriela Storkanova

MR Martin Reboun

HP Helena Poupetova

HJ Helena Jahnova

HH Helena Hulkova

JL Jana Ledvinova

LD Lenka Dvorakova

JS Jakub Sikora

MJ Milan Jirsa

MV Marie T. Vanier

MH Martin Hrebicek

This method is extracted from research article: Orphanet J Rare Dis, Apr 2020

Transcript, protein, metabolite and cellular studies in skin fibroblasts demonstrate variable pathogenic impacts of NPC1 mutations

DOI: 10.1186/s13023-020-01360-5

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Diagnostic filipin testing was performed at the time of individual diagnostic process as described in Vanier et al. [20] A main difference with the direct steady-state procedure was that dual culture slides (Lab-Tek chambers) with cells from each fibroblast line were first incubated in medium supplemented with 10% lipoprotein-deficient serum (LPDS) for 3 days, and then challenged for 24 h with (1) medium supplemented with LPDS and 50 μg/ml purified human LDL and (2) medium supplemented with 10% fresh total human serum, prior to filipin staining. A control and a typical NP-C cell lines were included in each diagnostic experiment. Fluorescence microscopic examination was performed using a selective DAPI filter and expert visual evaluation done as discussed by Vanier and Latour [18].

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