Name: Generatie en functionele verificatie van hypoxie-gevoelige chimere antigeenreceptor-T-cellen (Video) | JoVE Journal
Uploaded: 2024-06-14T14:00:00.000+00:00
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Description: Leer de generatie en functionele verificatie van hypoxie-gevoelige CAR-T-cellen voor kankerimmunotherapie. Dit protocol helpt onderzoekers bij het ontwikkelen van veiligere en effectievere celtherapieën.

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<meta charset="utf8"></head><body>Functionele validatie van hypoxie-gevoelige CAR-T-cellen voor selectieve tumoren

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<li>Liao, Q., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Engineering+T+cells+with+hypoxia-inducible+chimeric+antigen+receptor+%28HiCAR%29+for+selective+tumor+killing%5BTitle%5D)+AND+%22Biomark+Res%22%5BJournal%5D)">Engineering T cells with hypoxia-inducible chimeric antigen receptor (HiCAR) for selective tumor killing.</a> Biomark Res. 8 (1), 56(2020).</li>
<li>Ivan, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Hif%CE%B1+targeted+for+vhl-mediated+destruction+by+proline+hydroxylation%3A+Implications+for+o2+sensing%5BTitle%5D)+AND+%22Science%22%5BJournal%5D)">Hifα targeted for vhl-mediated destruction by proline hydroxylation: Implications for o2 sensing.</a> Science. 292 (5516), 464-468 (2001).</li>
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</ol>

<table><tbody><tr><td>1.5 mL Centrifuge tube</td><td>QSP</td><td>509-GRD-Q</td><td>Supernatants en cellen cellection Protocol Stap 2,3,4</td></tr><tr><td>10% ExpressCast PAGE</td><td>NCM biotech</td><td>P2012</td><td>Immunoblotting Protocol Stap 3</td></tr><tr><td>10x PBS</td><td>NCM biotech</td><td>20220812</td><td>Celcultuur Protocol Stap 4</td></tr><tr><td>10 mL pipet</td><td>Yueyibio</td><td>YB-25H</td><td>Pipetten Protocol Stap 1</td></tr><tr><td>10xTRIS-Glycine-SDS elektroforese buffer</td><td>Epizyme</td><td>3673020</td><td>Immunoblotting Protocol Stap 3</td></tr><tr><td>15 mL Centrifuge tube</td><td>Thermo Scientific</td><td>339650</td><td>Supernatants en cellen cellection Protocol Stap 1</td></tr><tr><td>25 cm2 EasYFlask</td><td>Thermo Scientific</td><td>156367</td><td>Celcultuur Protocol Stap 3,4</td></tr><tr><td>4x Protein SDS PAGE Loading Buffer</td><td>Takara</td><td>9173</td><td>Immunoblotting Protocol Stap 3</td></tr><tr><td>6-well flat-bottom weefselcultuur platen</td><td>Thermo Scientific</td><td>140675</td><td>T Cellen cultuur Protocol Stap 1</td></tr><tr><td>96-well zwarte flat-bottom weefselcultuur platen</td><td>Greiner</td><td>655090</td><td>Cytotoxiciteits test Protocol Stap 4</td></tr><tr><td>96-well ELISA platen</td><td>Corning</td><td>3590</td><td>ELISA Protocol Stap 5</td></tr><tr><td>96-well plate shaker</td><td>QILINBEIER</td><td>MH-2</td><td>Schudden Protocol Stap 4</td></tr><tr><td>96-well U-bottom weefselcultuur platen</td><td>Thermo Scientific</td><td>268200</td><td>Supernatants cellection Protocol Stap 4,5</td></tr><tr><td>anti-FLAG antilichaam</td><td>Sigma</td><td>F1804-50UG</td><td>Immunoblotting Protocol Stap 3</td></tr><tr><td>Carbinol</td><td>Sinopharm</td><td>10010061</td><td>Immunoblotting Protocol Stap 3</td></tr><tr><td>Koolstofdioxide incubator</td><td>Thermo Scientific</td><td>360</td><td>Celcultuur Protocol Stap 1,2,3,4</td></tr><tr><td>Celtelling plate</td><td>Hausser scientific</td><td>1492</td><td>Celtelling Protocol Stap 1,3,4</td></tr><tr><td>CELLection Pan Mouse IgG Kit</td><td>Thermo Scientific</td><td>11531D</td><td>Mouse IgG magnetische kralen Protocol Stap 1</td></tr><tr><td>Centrifuge</td><td>Thermo Scientific</td><td>75002432</td><td>Celcultuur Protocol Stap 1,3,4</td></tr><tr><td>Chemiluminescentie gel beeldvorming systeem</td><td>BIO-RAD</td><td>12003154</td><td>Immunoblotting Protocol Stap 3</td></tr><tr><td>Kobaltchloride oplossing (0.5 M)</td><td>bioleaper</td><td>BR4000203</td><td>Hypoxische conditie Protocol Stap 2,3,4</td></tr><tr><td>DMEM</td><td>Corning</td><td>10-103-CV</td><td>Celcultuur Protocol Stap 4</td></tr><tr><td>Elektronische weegschaal</td><td>Sartorius</td><td>PRACTUM612-1CN</td><td>Wegen Protocol Stap 5</td></tr><tr><td>FBS</td><td>BI</td><td>04-001-1ACS</td><td>Celcultuur Protocol Stap 3,4</td></tr><tr><td>GAPDH Mouse mAb</td><td>ABclonal</td><td>AC002</td><td>Immunoblotting Protocol Stap 3</td></tr><tr><td>Gel elektroforese apparaat</td><td>BIO-RAD</td><td>1645070</td><td>Immunoblotting Protocol Stap 3</td></tr><tr><td>GloMax Microplate Readers</td><td>Promega</td><td>GM3000</td><td>Luciferase activiteitsmeting Protocol Stap 4</td></tr><tr><td>Geit anti-Muis IgG (H+L)</td><td>Yeasen</td><td>P1126151</td><td>Immunoblotting Protocol Stap 3</td></tr><tr><td>Hoge snelheid microvriescentrifuge</td><td>eppendorf</td><td>5810&nbsp;R</td><td>Celcultuur Protocol Stap 1</td></tr><tr><td>Human IFN-&gamma; 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generatie en functionele verificatie van hypoxie-gevoelige chimere antigeenreceptor-t-cellen

Generation and Functional Verification of Hypoxia-Sensitive Chimeric Antigen Receptor-T Cells

Generatie en functionele verificatie van hypoxie-gevoelige chimere antigeenreceptor-T-cellen

Extensive studies have proven the promise of chimeric antigen receptor T (CAR-T) cell therapy in treating hematological malignancies. However, treating ...

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We beschrijven een kwantitatieve real-time in vitro cytolysis assay systeem om de potentie van chimere antigeen receptor T-cellen gericht op vloeibare en solide tumorcellen te evalueren. Dit protocol kan worden uitgebreid om andere immuuneffector cellen te beoordelen, evenals combinatie behandelingen.

Een real-time potentie test voor Chimerische antigeen receptor T-cellen gericht op vaste en hematologische kankercellen

Dit protocol beschrijft de methodologie voor het niet-invasief volgen van T-cellen die genetisch gemanipuleerd zijn om chimere antigeenreceptoren in vivo tot expressie te brengen met een klinisch beschikbaar platform.

Dynamische beeldvorming van chimere antigeenreceptor T-cellen met [18F]Tetrafluoroboraat positronemissietomografie/computertomografie

Dit protocol schetst het proces van het transduceren van primaire menselijke T-cellen met een gen van belang, waardoor compatibiliteit met Good Manufacturing Practice (GMP)-normen wordt gegarandeerd.

Een GMP-conforme procedure voor het genereren van gengemodificeerde T-cellen

Dit artikel beschrijft het productieproces voor chimere antigeenreceptor-T-cellen voor klinisch gebruik, met name met behulp van een geautomatiseerde celprocessor die virale transductie en kweek van T-cellen kan uitvoeren. We geven aanbevelingen en beschrijven valkuilen waarmee rekening moet worden gehouden tijdens de procesontwikkeling en implementatie van een klinische studie in een vroege fase.

Chimere antigeenreceptor T-celproductie op een geautomatiseerde celprocessor

Dit protocol stroomlijnt de productie van retrovirale vectoren en de transductie van T-cellen bij muizen, waardoor de efficiënte generatie van CAR-T-cellen van muizen wordt vergemakkelijkt.

Efficiënte generatie van muizen chimere antigeenreceptor (CAR)-T-cellen

Dit protocol biedt een gestroomlijnde workflow voor het genereren en testen van menselijke chimere antigeenreceptorregulerende T-cellen (CAR Tregs).

Generatie van menselijke chimere antigeenreceptorregulerende T-cellen

Dit protocol schetst een gedetailleerde procedure voor het genereren van voorbijgaande chimere antigeenreceptor (CAR) T-cellen met behulp van niet-integrerend mRNA voor kankerimmunotherapie en biedt betrouwbare methoden voor het evalueren van CAR-T-cellen en hun cytotoxische functie.

Een niet-virale benadering om voorbijgaande chimere antigeenreceptor-T-cellen te genereren met behulp van mRNA voor immunotherapie bij kanker

Aneuploïdie leidt tot instabiliteit van het genoom, die uiteindelijk celcyclus-gearresteerd cellen met complexe karyotypes produceert. Dit witboek biedt een eenvoudige en handige methode om te isoleren van de aneuploid cellen met complexe karyotypes dat niet langer verdelen.

Generatie en isolatie van de celcyclus-gearresteerd cellen met complexe Karyotypes

Leer de generatie en functionele verificatie van hypoxie-gevoelige CAR-T-cellen voor kankerimmunotherapie. Dit protocol helpt onderzoekers bij het ontwikkelen van veiligere en effectievere celtherapieën.

Generatie en functionele verificatie van hypoxie-gevoelige chimere antigeenreceptor-T-cellen

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