Beneficial effect of heat-induced antigen retrieval in immunocytochemical detection of intracellular antigens in alcohol-fixed cell samples

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61989592%3A15110%2F20%3A73591176" target="_blank" >RIV/61989592:15110/20:73591176 - isvavai.cz</a>

Výsledek na webu

<a href="https://journals.lww.com/appliedimmunohist/Abstract/2020/02000/Beneficial_Effect_of_Heat_induced_Antigen.11.aspx" target="_blank" >https://journals.lww.com/appliedimmunohist/Abstract/2020/02000/Beneficial_Effect_of_Heat_induced_Antigen.11.aspx</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1097/PAI.0000000000000689" target="_blank" >10.1097/PAI.0000000000000689</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Beneficial effect of heat-induced antigen retrieval in immunocytochemical detection of intracellular antigens in alcohol-fixed cell samples

Popis výsledku v původním jazyce

Immunohistochemistry (IHC) and immunocytochemistry (ICC) play an irreplaceable role in research and diagnostics. It is well known that antigen retrieval (AR) can, as a technique, have beneficial outcomes on IHC results when using formalin-fixed, paraffin-embedded tissue samples. The main purpose of AR is to break protein crosslinks which are formed during formalin fixation. Although AR was originally designed for FFPE samples, the usefulness of AR in ICC has been described. Cytological samples are often fixed in alcohol-based fixatives which does not lead to the formation of crosslinks. Therefore, alcohol-fixed samples can be successfully immunostained without AR. We investigated the effect of heat-induced antigen retrieval (HIAR) on alcohol-fixed HEK293 cell line samples and patient cytological samples from thyroid gland obtained by fine needle aspiration (FNA) technique. We compared indirect two-step ICC staining results performed according to the protocol with or without HIAR in citrate buffer pH6 for several antibodies. Utilising HIAR against intracellular antigens has beneficial effects. Therefore, more diluted antibodies can be used for satisfactory results. However, surface antigens were probably damaged by HIAR treatment. We demonstrated evident changes in cell surface topography after HIAR treatment by atomic force microscopy. Staining specificity of patient samples improves and background staining is reduced, allowing higher dilutions of primary antibody. Improving staining specificity is necessary for accurate diagnostics. Although we have shown the beneficial effect of HIAR for immunostaining intracellular antigens, proper staining protocol should be tested on appropriate controls for individual antibodies.

Název v anglickém jazyce

Beneficial effect of heat-induced antigen retrieval in immunocytochemical detection of intracellular antigens in alcohol-fixed cell samples

Popis výsledku anglicky

Immunohistochemistry (IHC) and immunocytochemistry (ICC) play an irreplaceable role in research and diagnostics. It is well known that antigen retrieval (AR) can, as a technique, have beneficial outcomes on IHC results when using formalin-fixed, paraffin-embedded tissue samples. The main purpose of AR is to break protein crosslinks which are formed during formalin fixation. Although AR was originally designed for FFPE samples, the usefulness of AR in ICC has been described. Cytological samples are often fixed in alcohol-based fixatives which does not lead to the formation of crosslinks. Therefore, alcohol-fixed samples can be successfully immunostained without AR. We investigated the effect of heat-induced antigen retrieval (HIAR) on alcohol-fixed HEK293 cell line samples and patient cytological samples from thyroid gland obtained by fine needle aspiration (FNA) technique. We compared indirect two-step ICC staining results performed according to the protocol with or without HIAR in citrate buffer pH6 for several antibodies. Utilising HIAR against intracellular antigens has beneficial effects. Therefore, more diluted antibodies can be used for satisfactory results. However, surface antigens were probably damaged by HIAR treatment. We demonstrated evident changes in cell surface topography after HIAR treatment by atomic force microscopy. Staining specificity of patient samples improves and background staining is reduced, allowing higher dilutions of primary antibody. Improving staining specificity is necessary for accurate diagnostics. Although we have shown the beneficial effect of HIAR for immunostaining intracellular antigens, proper staining protocol should be tested on appropriate controls for individual antibodies.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

30106 - Anatomy and morphology (plant science to be 1.6)

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2020

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

APPLIED IMMUNOHISTOCHEMISTRY &amp; MOLECULAR MORPHOLOGY

ISSN

1541-2016

e-ISSN

—

Svazek periodika

28

Číslo periodika v rámci svazku

2

Stát vydavatele periodika

US - Spojené státy americké

Počet stran výsledku

9

Strana od-do

166-174

Kód UT WoS článku

000516601200014

EID výsledku v databázi Scopus

2-s2.0-85052788339