Name: Procedures voor in vitro kweek van Treponema pallidum, de syfilisspirocheet (Video) | JoVE Journal
Uploaded: 2025-01-24T12:20:00.000+00:00
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Description: Leer de in vitro kweek van Treponema pallidum met Sf1Ep-cellen voor syfilisonderzoek. Deze methode maakt grootschalige productie en klonale culturen mogelijk zonder dierproeven.

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<meta charset="utf8"></head><body>Schaalbare in vitro kweek van T. pallidum met behulp van cocultuur van zoogdiercellen

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<li>Lin, M. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Longitudinal+TprK+profiling+of+in+vivo+and+in+vitro-propagated+Treponema+pallidum+subsp.+pallidum+reveals+accumulation+of+antigenic+variants+in+absence+of+immune+pressure%5BTitle%5D)+AND+%22PLoS+Negl+Trop+Dis%22%5BJournal%5D)">Longitudinal TprK profiling of in vivo and in vitro-propagated Treponema pallidum subsp. pallidum reveals accumulation of antigenic variants in absence of immune pressure.</a> PLoS Negl Trop Dis. 15 (9), e0009753(2021).</li>
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</ol>

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<table><tbody><tr><td>0.5 M EDTA, pH 8.0</td><td>Sigma&nbsp;</td><td>E8008</td><td></td></tr><tr><td>10x Earle&rsquo;s Balanced Salts, w/o Mg2+, Ca2+</td><td>Gibco</td><td>14155063</td><td></td></tr><tr><td>15 and 50 mL conische steriele wegwerp centrifuge buizen&nbsp;</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>2 mL cryogene vloeibare vials&nbsp;</td><td>Corning</td><td>430659</td><td></td></tr><tr><td>6-well cellcultuur platen voor T. pallidum cultivatie&nbsp;</td><td>Falcon</td><td>353046</td><td>De platen moeten beschikken over deksels met lage verdamping.</td></tr><tr><td>70% ethanol</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>75 cm2 weefselcultuur kolven met geventileerde doppen</td><td>Corning</td><td>43061U</td><td></td></tr><tr><td>93.5% stikstof, 5% CO2, en 1,5% zuurstof voor pre-equilibrering van medium en culturen</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>95% stikstof en 5% CO2 voor pre-equilibrering van medium en culturen</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>96-well laag verdamping heldere, platte bodem weefselcultuur behandelde microplates</td><td>Corning Falcon</td><td>353072</td><td></td></tr><tr><td>Verstelbare multi-kanaals pipet met 200 ul capaciteit&nbsp;</td><td>N/A</td><td>N/A</td><td>Optioneel, maar zeer nuttig voor klonen</td></tr><tr><td>Cellcultuur kwaliteit water</td><td>Sigma</td><td>W3500</td><td></td></tr><tr><td>CMRL 1066 zonder L-Glutamine of Fenol Rood</td><td>US Biological</td><td>C5900-03A</td><td></td></tr><tr><td>CO2 voor tri-gas en weefselcultuur incubators&nbsp;</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>Cryogene vloeibare stikstof weefselcultuur opslagtank</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>D-glucose</td><td>Sigma-Aldrich</td><td>G6152</td><td></td></tr><tr><td>Weggebruikbare filter units, 0,2 &micro;m, > 100 mL capaciteit</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>Weggebruikbare pipetten: 25 mL, 10 mL, 5 mL, aspirant</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>DL-Dithiothreitol</td><td>Sigma-Aldrich</td><td>D9779</td><td></td></tr><tr><td>D-Mannitol&nbsp;</td><td>Sigma-Aldrich</td><td>M1902</td><td></td></tr><tr><td>DMSO (steriele cellcultuur kwaliteit)</td><td>Sigma-Aldrich</td><td>D2650</td><td></td></tr><tr><td>Eagle&rsquo;s MEM</td><td>Sigma-Aldrich</td><td>M4655</td><td></td></tr><tr><td>Foetaal bovien serum, hitte geactiveerd&nbsp;</td><td>Sigma-Aldrich</td><td>F4135</td><td>We bevelen dit product sterk aan. Moet pre-screenen voor T. pallidum cultuur compatibiliteit als u een ander merk of catalogusnummer gebruikt.</td></tr><tr><td>Freezer met mogelijkheid om -70 &deg;C of -80 &deg;C te handhaven</td><td>N/A</td><td>N/A</td><td>Voor opslag van T. pallidum; vloeibare stikstof opslag kan ook worden gebruikt</td></tr><tr><td>Freezing medium (Sf1Ep medium + 10% [v/v] DMSO)</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>Gasflessen met geschikte fittingen</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>GasPak 150 geventileerde anaërobe pot (Brewer Pot)</td><td>Fisher Scientific</td><td>11-816</td><td></td></tr><tr><td>Glycerol</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>Helber tellkamers met Thoma rulings</td><td>Hawksley Medical and Laboratory Equipment</td><td></td><td>Voor het kwantificeren van T. pallidum</td></tr><tr><td>Hemocytometer</td><td>N/A</td><td>N/A</td><td>&nbsp;Voor Sf1Ep cel kwantificering</td></tr><tr><td>Incubator tank schakelaar</td><td>NuAire&nbsp;</td><td>NU-1550 TankGuard Automatic CO2 Incubator Tank Switch</td><td>Optioneel, maar zeer nuttig bij het handhaven van geschikte O2 omstandigheden.</td></tr><tr><td>Inverteer microscoop met fase contrast optiek</td><td>N/A</td><td>N/A</td><td>Voor het bekijken van Sf1Ep cel culturen</td></tr><tr><td>L-Glutamine</td><td>Sigma-Aldrich</td><td>G7513</td><td></td></tr><tr><td>L-Histidine&nbsp;</td><td>Sigma-Aldrich</td><td>H6034</td><td></td></tr><tr><td>MEM Niet-Essentiële Aminozuren</td><td>Gibco</td><td>11140-050</td><td></td></tr><tr><td>Microscoop met darkfield condensor</td><td>N/A</td><td>N/A</td><td>De microscoop moet een 40x objectief en 15x oogstukken hebben.</td></tr><tr><td>MOPS</td><td>Sigma-Aldrich</td><td>M3183</td><td></td></tr><tr><td>Multi-kanaals adapter voor aspirator&nbsp;</td><td>Integra</td><td>155520</td><td>Optioneel, maar nuttig voor klonen</td></tr><tr><td>NaHCO3 (7,5%)</td><td>Sigma-Aldrich</td><td>S8761</td><td></td></tr><tr><td>Stikstof voor tri-gas incubator</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>Resazurin</td><td>Sigma-Aldrich</td><td>R7017</td><td></td></tr><tr><td>Sf1Ep (NBL-11) cellen</td><td>American Type Culture Collection</td><td>CCL-68</td><td></td></tr><tr><td>Natriumpyruvaat</td><td>Sigma-Aldrich</td><td>S8636</td><td></td></tr><tr><td>Steriele PBS (zonder calciumchloride en magnesiumchloride)&nbsp;</td><td>Sigma-Aldrich</td><td>D8537</td><td></td></tr><tr><td>Steriele reagent reservoirs, 50 of 100 mL grootte</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>T. pallidum monster, bevroren of vers&nbsp;</td><td></td><td></td><td>van een konijn infectie of in vitro cultuur</td></tr><tr><td>Weefselcultuur incubator gehandhaafd bij 37 &deg;C, 5% CO2</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>Tri-gas weefselcultuur incubator gehandhaafd bij 34 &deg;C, 5% CO2, 1,5% O2</td><td>Thermofisher</td><td>Heracell&trade; VIOS 160i Tri-Gas CO2 Incubator</td><td>Optioneel; anaërobe potten kunnen in plaats daarvan worden gebruikt (zie Ref. 17)</td></tr><tr><td>Trypsin-EDTA oplossing&nbsp;</td><td>Sigma-Aldrich</td><td>T4049</td><td></td></tr><tr><td>Vacuum bron (bijv. huis vacuüm), vacuümbuis, vacuümmeter en connectoren</td><td>N/A</td><td>N/A</td><td></td></tr><tr><td>Water, geschikt voor celcultuur, filter

procedures voor in vitro kweek van treponema pallidum, de syfilisspirocheet

Procedures for In Vitro Cultivation of Treponema pallidum, the Syphilis Spirochete

Procedures voor in vitro kweek van Treponema pallidum, de syfilisspirocheet

For over a century, Treponema pallidum subsp. pallidum, the spiral-shaped bacterium that causes syphilis, could only be propagated by inoculation and ...

procedures-for-vitro-cultivation-treponema-pallidum-syphilis

Research

JoVE Journal

Immunology and Infection

Immunologie en Infectie

Source: Eggers, C. H. Phage-Mediated Genetic Manipulation of the Lyme Disease Spirochete Borrelia burgdorferi. J. Vis. Exp. (2022)This video demonstrates the preparation of a genetically modified Borrelia burgdorferi culture engineered with dormant bacteriophage DNA carrying an antibiotic resistance gene. Following antibiotic selection and incubation, a chemical inducer triggers DNA damage, leading to excision, replication, and assembly of new virions containing the resistance gene. After a...

Preparation and Induction of Borrelia burgdorferi Cultures for Viral Particle Production

Source: Berthold, A., et al. Cultivation Methods of Spirochetes from Borrelia burgdorferi Sensu Lato Complex and Relapsing Fever Borrelia. J. Vis. Exp.(2022)This video demonstrates the cultivation of pathogenic spirochetes from infected skin tissue to model dermal infection. It highlights their localization in the dermis, interaction with the extracellular matrix, and survival during selective culture conditions.

Cultivation of Pathogenic Spirochetes to Investigate Skin Infection

Snelle en efficiënte kwantificering van intracellulaire M. tuberculosis-groei is cruciaal voor het nastreven van verbeterde therapieën tegen tuberculose (TB). Dit protocol beschrijft een op bouillon gebaseerde colorimetrische detectietest met behulp van een geautomatiseerd vloeistofkweeksysteem om de Mtb-groei te kwantificeren in macrofagen die worden behandeld met kandidaat-gastheergerichte therapieën.

Een geautomatiseerd kweeksysteem voor gebruik bij preklinische tests van gastheergerichte therapieën voor tuberculose

Vanwege de ondoorzichtigheid van de bodem kunnen interacties tussen de samenstellende microben niet gemakkelijk worden gevisualiseerd met cellulaire resolutie. Hier worden twee microfluïdische hulpmiddelen gepresenteerd, die nieuwe mogelijkheden bieden voor het onderzoeken van schimmel-microbiële interacties. De apparaten zijn veelzijdig en eenvoudig te gebruiken, waardoor hoge spatiotemporale controle en beeldvorming met hoge resolutie op cellulair niveau mogelijk is.

Microfluïdische hulpmiddelen voor het onderzoeken van schimmel-microbiële interacties op cellulair niveau

Het vermogen van bacteriofaag om DNA tussen bacteriële cellen te verplaatsen, maakt ze effectieve hulpmiddelen voor de genetische manipulatie van hun bacteriële gastheren. Hier wordt een methodologie gepresenteerd voor het induceren, herstellen en gebruiken van φBB-1, een bacteriofaag van Borrelia burgdorferi, om heterologe DNA te transduceren tussen verschillende stammen van de ziekte van Lyme spirocheet.

Faag-gemedieerde genetische manipulatie van de ziekte van Lyme Spirochete Borrelia burgdorferi

In vitro kweek is een directe detectiemethode voor de aanwezigheid van levende bacteriën. Dit protocol beschrijft methoden voor de kweek van diverse Borrelia spirocheten, waaronder die van het Borrelia burgdorferi sensu lato-complex, recidiverende koorts Borrelia-soorten en Borrelia miyamotoi.  Deze soorten zijn kieskeurig en langzaam groeiend, maar kunnen worden gekweekt.

Teeltmethoden van Spirocheten uit Borrelia burgdorferi Sensu Lato Complex en Relapsing Fever Borrelia

Elektroporatie is een snelle, breed toegepaste methode voor het introduceren van exogene DNA in het geslacht Rickettsia. Dit protocol biedt een bruikbare elektroporatiemethode voor de transformatie van obligate intracellulaire bacteriën in het geslacht Rickettsia.

Een elektroporatiemethode om Rickettsia spp. te transformeren met een fluorescerende eiwitexpressie shuttlevector in tekencellijnen

Deze studie toont een gedetailleerd protocol voor het uitvoeren van ultrastructuurexpansiemicroscopie in drie in vitro levenscyclusstadia van Trypanosoma cruzi, de ziekteverwekker die verantwoordelijk is voor de ziekte van Chagas. We nemen de geoptimaliseerde techniek voor cytoskeletale eiwitten en pan-proteoomlabeling op.

Ultrastructurele expansiemicroscopie in drie in vitro levenscyclusstadia van Trypanosoma cruzi

Dit artikel beschrijft de procedure voor de vorming en visualisatie van een bacteriële biofilm gegroeid binnen een 8-well kamer...

In vitro biofilmvorming in een 8-well Kamer Dia

Cercarial Transformatie en In vitro Teelt van Schistosoma mansoni Schistosomules

Leer de in vitro kweek van Treponema pallidum met Sf1Ep-cellen voor syfilisonderzoek. Deze methode maakt grootschalige productie en klonale culturen mogelijk zonder dierproeven.

Procedures voor in vitro kweek van Treponema pallidum, de syfilisspirocheet

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