Name: Isolamento a nucleo singolo dal tessuto endorerectomico coronarico di pazienti con bypass coronarico (Video) | JoVE Journal
Uploaded: 2026-04-03T13:08:00.000+00:00
Duration: 9 min 9 s
Description: Scopri un protocollo per isolare nuclei intatti da tessuto endarterectomico coronarico umano. Questo metodo consente il profilo trascrittomico di placche aterosclerotiche ad alto rischio tramite snRNA-seq.

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<li>Bakken, T. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Single-nucleus+and+single-cell+transcriptomes+compared+in+matched+cortical+cell+types%5BTitle%5D)+AND+%22PLoS+One%22%5BJournal%5D)">Single-nucleus and single-cell transcriptomes compared in matched cortical cell types.</a> PLoS One. 13 (12), e0209648(2018).</li>
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<li>Hu, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Single-nucleus+transcriptomic+survey+of+cell+diversity+and+functional+maturation+in+postnatal+mammalian+hearts%5BTitle%5D)+AND+%22Genes+Dev%22%5BJournal%5D)">Single-nucleus transcriptomic survey of cell diversity and functional maturation in postnatal mammalian hearts.</a> Genes Dev. 32 (19-20), 1344-1357 (2018).</li>
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</ol>

<table><tbody><tr><td>100 & mu; Filtro m</td><td>Miltenyi Biotech&nbsp;</td><td>130-098-463</td><td></td></tr><tr><td>30 & mu; Filtro m</td><td>Miltenyi Biotech&nbsp;</td><td>130-098-458</td><td></td></tr><tr><td>Albumina Sirica Bovina</td><td>HiMedia</td><td>TC546-25G</td><td></td></tr><tr><td>Centrifuga</td><td>MPW</td><td>NA</td><td>https://mpw.pl/en/offer/mpw-352r</td></tr><tr><td>Microscopio composto</td><td>Nikon</td><td>NA&nbsp;</td><td>Microscopio rovesciato Nikon Eclipse Ts2 (https://www.microscope.healthcare.nikon.com/en_AOM/products/inverted-microscopes/eclipse-ts2)</td></tr><tr><td>Microscopio Confocale &nbsp;</td><td>Nikon</td><td>NA</td><td>Microscopio confocale rovesciato Nikon Ti2-E (https://www.microscope.healthcare.nikon.com/en_AOM/products/inverted-microscopes/eclipse-ti2-series)</td></tr><tr><td>Copertine</td><td>HiMedia</td><td>BG011C</td><td></td></tr><tr><td>Fiala di campione CRYOCHIL a bocca larga</td><td>Tarsons</td><td>883191</td><td></td></tr><tr><td>DAPI</td><td>Sigma</td><td>D9542-5MG</td><td></td></tr><tr><td>DPBS</td><td>Gibco (termofischer)</td><td>10010-023</td><td></td></tr><tr><td>Tubo centrifuga (15 mL)</td><td>Tarsons</td><td>546021</td><td></td></tr><tr><td>Citometro a flusso&nbsp;</td><td>BD in Biologia</td><td>663029</td><td>BD FACSLyric (3L12C Instrument CEIVD)</td></tr><tr><td>Emocitometro</td><td>Axibio</td><td>CCB 100</td><td></td></tr><tr><td>Tubo microcentrifuga (1,5 mL)</td><td>Tarsons</td><td>500010C</td><td></td></tr><tr><td>Tubo microcentrifugo (5 mL)</td><td>Tarsons</td><td>500050</td><td></td></tr><tr><td>MgCl2</td><td>Sigma</td><td>M1028-100ML</td><td></td></tr><tr><td>Micro pelistino</td><td>Tarsons</td><td>160020</td><td></td></tr><tr><td>NaCl</td><td>Sigma</td><td>S9888-500G</td><td></td></tr><tr><td>NP-40</td><td>HiMedia</td><td>MB143-100ML</td><td></td></tr><tr><td>Acqua priva di nucleasi</td><td>Gibco (termofischer)</td><td>W4502-1L</td><td></td></tr><tr><td>Inibitore RNasi</td><td>Sigma</td><td>333540201</td><td></td></tr><tr><td>RPMI 1640</td><td>Gibco (termofischer)</td><td>11875093</td><td></td></tr><tr><td>Diapositiva&nbsp;</td><td>Riviera</td><td>72910135</td><td></td></tr><tr><td>Tris-HCl</td><td>Sigma</td><td>T2194-100ML</td><td></td></tr><tr><td>Trippan blu & nbsp;</td><td>Sigma</td><td>302643-25G</td><td></td></tr></tbody></table>

isolamento a nucleo singolo dal tessuto endorerectomico coronarico di pazienti con bypass coronarico

Single Nuclei Isolation from Coronary Endarterectomy Tissue of Coronary Artery Bypass Graft Patients

Il tessuto endarterectomico coronarico umano è stato raccolto da pazienti sottoposti a interventi di chirurgia CABG. Il tessuto è stato pesato e processato per l'isolamento dei nuclei, e ulteriori nuclei isolati sono stati confermati tramite citometria a flusso e microscopia confocale (Figura 1).
Valutazione della qualità dell'isolamento dei nuclei con il blu di tripano Il tessuto tenuto in mezzo RPMI è stato pesato e ha misurato 247,8 mg. Il t...

Isolamento a nucleo singolo dal tessuto endorerectomico coronarico di pazienti con bypass coronarico

Atherosclerosis is a chronic, progressive inflammatory condition characterized by the accumulation of plaque in the walls of arteries. During the ...

single-nuclei-isolation-from-coronary-endarterectomy-tissue-coronary

Research

Ricerca

JoVE Journal

Biology

Biologia

Nuclei cardiaca sono isolati tramite sedimentazione densità e immunolabeled con anticorpi contro materiale pericentriolar 1 (PCM-1) per identificare e ordinare nuclei cardiomiociti mediante citometria a flusso.

Isolamento dei Nuclei Cardiomyocyte dal post-mortem del tessuto

Le attuali conoscenze sulla base cellulare di malattie cardiache si basa principalmente su studi su modelli animali. Qui descriviamo e validare un nuovo metodo per ottenere singoli cardiomiociti vitali da piccoli campioni chirurgici di miocardio ventricolare umana. Miociti ventricolari umani possono essere utilizzati per studi elettrofisiologici e test della droga.

Isolamento e caratterizzazione funzionale di Human ventricolare Cardiomyocytes da campioni freschi chirurgici

La capacità di isolare le cellule endoteliali delle valvole cardiache (VEC) è fondamentale per comprendere i meccanismi di sviluppo, mantenimento e malattia delle valvole.  Qui descriviamo l'isolamento di VEC da topi Tie2-GFP embrionali e adulti utilizzando FACS che consentiranno studi che determineranno il contributo dei VEC nei processi di sviluppo e malattia.

Isolamento di cellule murine endoteliali Valve

Lo scopo di questo protocollo è quello di dimostrare l'isolamento e coltura tecniche di cellule muscolari murine primarie lisce vascolari (VSMC) dalla circolazione coronarica. Una volta VSMC sono stati isolati, essi possono essere utilizzati per vari tecniche di coltura standard.

L&#39;isolamento di Murine coronarica vascolari cellule muscolari lisce

Questo protocollo è preparato per condividere il nostro metodo di isolamento delle cellule endoteliali coronariche di topo a scopo di imaging o per condurre esperimenti di biologia molecolare.

Isolamento di cellule di topo coronarica endoteliali

tessuto cardiaco umano alberga popolazioni di cellule precursore perivascolari multipotenti che possono essere adatti per la rigenerazione del miocardio. La tecnica qui descritta permette l'isolamento e la purificazione simultanea di due popolazioni di cellule stromali multipotenti associati con i vasi sanguigni nativi, cioè CD146 + CD34 - periciti e CD34 + CD146 - cellule avventiziale, dal miocardio umano.

Isolamento di perivascolare Multipotent precursori delle cellule popolazioni dal cardiache umane Tissue

Cellule endoteliali tumorali sono fattori determinanti importanti del microambiente tumorale e il decorso della malattia. Qui, un protocollo per l'isolamento di cellule endoteliali pure e praticabile da carcinoma colorettale umano e colon normale a essere impiegata nella ricerca di droga test e la patogenesi è descritto.

Isolamento di cellule endoteliali umane da Colon normale e Carcinoma colorettale - un protocollo migliorato

L'obiettivo di questo lavoro è quello di sviluppare un metodo per isolare in modo riproducibile i cardiomiociti dal cuore adulto e misurare il contenuto di DNA e la nucleazione.

Isolamento dei cardiomiociti da cuori fissi per l'immunocitochimica e l'analisi dello ploidy

Vi presentiamo un protocollo per l'isolamento delle cellule endocardiali e coronariche dai cuori dei ratti attraverso la digestione sequenziale dei tessuti in un buffer di digestione, la raccolta cellulare da cicli di centrifuga ricorrenti e la purificazione cellulare utilizzando microsfere CD31 anti-rat.

Isolamento delle cellule endocartali e coronarie dal muro libero ventricolare del cuore di ratto

Qui, presentiamo un protocollo che descrive la preparazione dei nuclei cellulari. Dopo la microdissezione e la dissociazione enzimatica del tessuto cardiaco in singole cellule, le cellule progenitrici sono state congelate, seguite dall'isolamento di cellule vitali pure, che sono state utilizzate per il sequenziamento dell'RNA a singolo nucleo e dal test a nucleo singolo per la cromatina accessibile alla trasposasi con analisi di sequenziamento ad alto rendimento.

Isolamento di nuclei da cellule progenitrici cardiache di topo per il profilo dell'epigenoma e dell'espressione genica a risoluzione di singole cellule

Questo studio presenta un modello suino chirurgico di ischemia miocardica cronica dovuta a stenosi progressiva dell'arteria coronaria, con conseguente compromissione della funzione cardiaca senza infarto. Dopo l'ischemia, gli animali vengono sottoposti a trapianto di bypass aortocoronarico off-pump con posizionamento epicardico di cerotti di collagene carichi di esosomi derivati da cellule staminali. Questa terapia aggiuntiva migliora la funzione miocardica e il recupero.

Modello suino chirurgico di ischemia miocardica cronica trattata con cerotto di collagene carico di esosomi e innesto di bypass aortocoronarico off-pump

Scopri un protocollo per isolare nuclei intatti da tessuto endarterectomico coronarico umano. Questo metodo consente il profilo trascrittomico di placche aterosclerotiche ad alto rischio tramite snRNA-seq.

Isolamento a nucleo singolo dal tessuto endorerectomico coronarico di pazienti con bypass coronarico

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