Name: Automated Cell Culture System Settings for Maintenance and Differentiation of Human Pluripotent Stem Cells
Uploaded: 2024-01-26T13:30:00
Description: Watch this Scientific Journal Video about Automated Cell Culture System Settings for Maintenance and Differentiation of Human Pluripotent Stem Cells at JoVE.com

automated cell culture system settings for maintenance and differentiation of human pluripotent stem cells

Induceret pluripotent stamcelledifferentiering ved hjælp af et automatiseret dyrkningssystem

This article aims to provide actual and detailed handling procedures for an automated culture system for human induced pluripotent stem cells (iPSC), which we produced by collaborating with a company, and to show representative results.
Since the publication of the article in 2007, iPSC has been attracting attention all over the world1. Due to its greatest feature of being able to differentiate into any type of somatic cell, it is expected to be applied in various fields such as regenerative medicine, elucidating the causes of intractable diseases, and developing new therapeutic drugs2,3. In addition, using human iPSC-derived somatic cells could reduce animal experiments, which are subject to significant ethical restrictions. Although numerous homogeneous iPSCs are constantly required to research new methods with iPSCs, it is too laborious to manage them. Moreover, handling iPSC is difficult because of its high sensitivity, even to subtle cultural and environmental changes.
To solve this problem, automated culture systems are expected to perform tasks instead of humans. Some groups have developed a few automated human pluripotent stem cell culture systems for cell maintenance and differentiation and published their achievements4,5,6. These systems equip multiarticulated robotic arm(s). Robotic arms have not only merit in that they highly mimic human arm movements but also demerit in that they require higher cost(s) for the arm(s), larger and heavier system packaging, and time-consuming education efforts by the engineers to obtain the aimed movements7,8. In order to make it easier to introduce the apparatus to more research facilities at the points of economic, space, and human resource consumption, we have developed a novel automated culture system for the maintenance and differentiation of iPSC into various cell types9.
Our rationale for the new system was to adopt an X-Y-Z axis rail system instead of multiarticulated robotic arms9. To replace the complex hand-like functions of robotic arms, we applied a new idea to this system, which can automatically change three types of specific functional arm tips. Here, we also indicate how users can easily make task schedules with simple orders on software because of the lack of requirements for engineers&#39; contributions throughout the process.
One of the robotic culture systems has demonstrated the making of embryoid bodies using 96-well plates as 3D cell aggregates for differentiation4. The system reported here cannot handle 96-well plates. One achieved the current good manufacturing practice (cGMP) grade using a cell line, although it was not a human pluripotent stem cell5. The automated culture system detailed here has now been developed with the specific aim of helping laboratory experiments (Figure 1). However, it has enough systems to keep clean levels equivalent to a level IV safety cabinet.

Forfatter Spotlight: Automatisering af iPSC-kultur for forbedret reproducerbarhed 

Et automatiseret dyrkningssystem til vedligeholdelse og differentiering af menneskeskabte pluripotente stamceller

This study focuses on automating the culture of iPS cells. Our goal is to reduce the variability due to manual experimentations and human labor, by utilizing smaller and more affordable equipment, which leads to enhance the reproducibility, and provide opportunities for more researchers who are even unfamiliar with iPS cells. Several types of automated culture machines have emerged, but most of them are still large, costly, and can only perform partial tasks.The simpler structure of the working urn has reduced the size and cost of the equipment. In addition, the tasks from cell maintenance to differentiation induction can be performed automatically, with only prior material preparation, and task setting on software. Machine repetition is remarkably accurate compared to human repetition.The use of such machines not only improves repeat usability, but also reduce human labor. Furthermore, if the parameters in each protocol are shared, researchers unfamiliar with iPS cells will be able to achieve similar results as well-experienced researchers, making it possible for them to easily enter into research using iPS cells. We are exploring the possibility of incorporating an AI-based image judgment system to automate the selection of the next work schedule, setting date, and work content from the learning of the experiment results.We also anticipate that allowing researchers to tailor their working protocols to their preferences, and share them online, will not only enhance reproducibility among the world researchers, but also encourage global research collaboration.

Watch this Scientific Journal Video about Automated Cell Culture System Settings for Maintenance and Differentiation of Human Pluripotent Stem Cells at JoVE.com

Automated Cell Culture System Settings for Maintenance and Differentiation of Human Pluripotent Stem Cells  

Automatiserede cellekultursystemindstillinger til vedligeholdelse og differentiering af humane pluripotente stamceller

For at begynde at indlæse skal du tænde for det automatiserede cellekulturinstrument og starte operativsystemet. Klik på indlæsningsknappen på softwaren, efterfulgt af start af indlæsningsforberedelse. Placer nu opvasken i apparatet i deres specifikke positioner.Luk det forreste skydevindue, og tryk på knappen til bekræftelse af mekanisk sikkerhed. Vælg typen og mængden af retter i softwaren. Klik derefter på indlæsningsforberedelse afsluttet, efterfulgt af startknappen for indlæsning.Når skålen er uploadet til systemet, skal du registrere noter om hver enkelt parabol i softwaren. Klik derefter på registrering for at afslutte indlæsningen. For at aflæse opvasken skal du først klikke på aflæsning.Vælg derefter de retter, der skal fjernes. Tryk derefter på startknappen for losningsforberedelsen, og klik på aflæsningsstart. Når instrumentet overfører opvasken fra inkubatoren til arbejdsbordet, skal du klikke på skålens fjernelsesknap.Åbn det forreste skydevindue manuelt og fjern opvasken. Luk straks det forreste skydevindue, og tryk på sikkerhedsbekræftelsesknappen for at fuldføre aflæsningen. For at genopfylde forbrugsstoffer skal du klikke på knappen forbrugsstoffer på softwaren.For pipetterne skal du klikke på pipetteknappen efterfulgt af genopfyldningsknappen. Vælg det stativ, der skal genopfyldes. Tryk derefter på genopfyld start.Åbn vinduet med skydestativet manuelt, og genopfyld pipetterne efter behov. Luk derefter vinduet og tryk på den mekaniske sikkerhedsknap. Klik på knappen genopfyldning afsluttet efterfulgt af genopfyldningsopsætningen for at indtaste oplysningerne.Klik derefter på registreringsknappen og knappen genopfyld afsluttet for at afslutte. På samme måde genopfyldes rørene og medierne efter ønske. Klik derefter på knappen Luk for at afslutte genopfyldningsopsætningen.For at vælge opgaver skal du klikke på opgaveknappen på softwaren. Tryk nu på opgaveindstillingsknappen, og vælg den ønskede opgave fra opgavelisten. Klik derefter på knappen næste trin.Indtast dato og klokkeslæt for den opgave, der skal udføres. Klik derefter på registreringsknappen. Vælg nu skålen eller pladen for at udføre opgaven.Klik derefter på registreringsknappen igen, bekræft den valgte opgave igen, og tryk på registrering. Klik derefter på start for automatisk at starte opgaven på det angivne tidspunkt og dato. For at annullere en planlagt opgave på forhånd skal du klikke på stop.Vælg derefter den opgave, der skal afbrydes. Klik på rediger opgave, efterfulgt af knappen Annuller opgave, og bekræft sletningen af opgaven. For at udføre cellepasning og differentiering skal du vælge de ønskede kulturindstillinger i henhold til den ønskede celle, der skal opnås.Den automatiserede cellekultur tillod jævn celleproliferation over tid. Immunocytometrisk analyse viste, at de inducerede pluripotente stamceller forblev udifferentierede. Dette blev bekræftet af immunhistokemiske analyser.Der blev ikke observeret alleliske abnormiteter efter fem vedligeholdelseskulturer. De pluripotente stamceller blev observeret at gennemgå differentiering i kardiomyocytter, hepatocytter, neuronale forløberceller og keratinocytter med det automatiserede system.