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Samenvatting - Celcultuur en immunochemie
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Samenvatting Celcultuur en Immunochemie - 2de bachelor Biomedische Wetenschappen
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, Celcultuur en immunochemie | [Author Name]Table of Contents
HS01: inleiding ................................................................................................................................................................................. 5
1. Geschiedenis ...............................................................................................................................................5
1.1. Drie belangrijke ontdekkingen (basis celcultuur) ................................................................................................................................ 5
1.2. Poliovirus en -vaccin .................................................................................................................................................................................. 5
2. Voordelen van weefselkweekculturen .........................................................................................................6
2.1. Controle omgeving .................................................................................................................................................................................... 6
2.2. Goede karakterisatie en homogeniteit ................................................................................................................................................ 6
2.3. Kostprijs ....................................................................................................................................................................................................... 6
2.4. In vitro modellering van in vivo omstandigheden ............................................................................................................................... 6
3. Nadelen / beperkingen van weefselculturen ...............................................................................................6
3.1. Expertise...................................................................................................................................................................................................... 6
3.2. Hoeveelheid................................................................................................................................................................................................ 6
3.3. Instabiliteit ................................................................................................................................................................................................... 6
3.4. Cultuuromgeving vs. in vivo ..................................................................................................................................................................... 6
HS02: cel- en weefselkweekculturen: definities ....................................................................................................................... 7
1. Soorten cultuurmethoden (6) .......................................................................................................................7
1.1. Orgaanculturen .......................................................................................................................................................................................... 7
1.2 Primaire culturen ........................................................................................................................................................................................ 7
1.3. Expant cultuur ............................................................................................................................................................................................. 7
1.4. Eindige celculturen ..................................................................................................................................................................................... 8
1.5. Continue cellijnen = geïmmortaliseerde cellijnen ............................................................................................................................... 8
1.6. Organotypische celculturen ..................................................................................................................................................................... 8
Cel- en weefselculturen ..........................................................................................................................................9
2. Overzicht grafiek .........................................................................................................................................9
HS03: soorten cellen en culturen............................................................................................................................................... 10
1. Verkrijgen van cellen voor celkweek .........................................................................................................10
1.1. Migratie uit het weefsel ........................................................................................................................................................................ 10
1.2. Mechanische dissociatie ........................................................................................................................................................................ 10
1.3. Enzymatische dissociatie ....................................................................................................................................................................... 10
2. Celadhesie .................................................................................................................................................11
2.1. Celadhesiemoleculen (CAM’s) .............................................................................................................................................................. 11
2.2. Cel-ECM-adhesie .................................................................................................................................................................................... 12
2.3. Cel-cel-adhesie ....................................................................................................................................................................................... 12
3. Soorten cellen in kweek .............................................................................................................................12
3.1. Stamcellen ................................................................................................................................................................................................ 12
4.2. Kankercellen .................................................................................................................................................................................................. 13
4. Immortalisatie van cellen ..........................................................................................................................13
4.1. Transformatie .......................................................................................................................................................................................... 13
4.2. Voor- en nadelen ................................................................................................................................................................................... 14
5. Selectie van celtype ...................................................................................................................................14
6. Celstocks ....................................................................................................................................................14
7. Celbanken..................................................................................................................................................14
MRC-5 cellen.......................................................................................................................................................................................................... 14
HeLa cellen ............................................................................................................................................................................................................. 14
HS04: Subcultivatie ...................................................................................................................................................................... 15
1. Inleiding.....................................................................................................................................................15
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, Celcultuur en immunochemie | [Author Name]
2. Cellen oogsten: (2) manieren.....................................................................................................................16
2.3. Enzymatisch losmaken ............................................................................................................................................................................ 16
2.4. Mechanisch losmaken ............................................................................................................................................................................. 16
3. Cellen tellen ...............................................................................................................................................16
3.3. Tellen ......................................................................................................................................................................................................... 16
3.4. Bürker telkamer ...................................................................................................................................................................................... 16
4. Cellen uitzaaien .........................................................................................................................................17
HS05: Cryopreservatie van dierlijke cellen............................................................................................................................. 18
1. Het maken van een celbank ......................................................................................................................18
1.1. Problemen ................................................................................................................................................................................................ 18
1.2. Oplossing: cryopreservatie .................................................................................................................................................................. 18
2. Principes cryopreservatie ...........................................................................................................................18
2.1. Mechanisme van invriezen van cellen ................................................................................................................................................ 18
2.2. Celdichtheid ............................................................................................................................................................................................. 19
2.3. Invriesmedium .......................................................................................................................................................................................... 19
3. Invriesmaterialen .............................................................................................................................................19
3.1. Invriesampullen .............................................................................................................................................................................................. 19
3.2. Opslag van ampullen ............................................................................................................................................................................ 20
4. Ontdooien van een cryostock ....................................................................................................................20
4.1. Ontdooitmethode.......................................................................................................................................................................................... 20
4.2. Vitrificatie ....................................................................................................................................................................................................... 20
HS06: Materialen en media ........................................................................................................................................................ 21
1. Recipiënten ................................................................................................................................................21
1.1. Materiaal ................................................................................................................................................................................................. 21
1.2. Soorten cultuurflessen / platen............................................................................................................................................................ 21
1.3. Gespecialiseerde systemen.................................................................................................................................................................. 22
2. Media ..............................................................................................................................................................25
2.1. Verschillende media ..................................................................................................................................................................................... 25
2.2. Basisingrediënten.................................................................................................................................................................................... 25
2.3. Extra ingrediënten.................................................................................................................................................................................. 25
2.4. Serum-vrij medium .................................................................................................................................................................................. 27
2.5. pH .............................................................................................................................................................................................................. 27
2.6. Osmoraliteit ............................................................................................................................................................................................. 28
3. Behandeling kweekoppervlak: coating ......................................................................................................28
3.1. Doel ........................................................................................................................................................................................................... 28
3.2. Werkwijze................................................................................................................................................................................................ 28
Hoofdstuk 7: de celcultuuromgeving ........................................................................................................................................ 29
1. Inleiding.....................................................................................................................................................29
2. 𝑪𝑶𝟐-incubatoren .......................................................................................................................................29
2.1. Types ......................................................................................................................................................................................................... 29
2.2. Temperatuur ............................................................................................................................................................................................ 29
2.3. 𝑪𝑶𝟐 .......................................................................................................................................................................................................... 29
2.4. Vochtigheid .............................................................................................................................................................................................. 29
2.5. Desinfectie en sterilisatie ...................................................................................................................................................................... 29
3. LAF-kast = laminaire luchtstroomkast .......................................................................................................30
3.1. Functie ....................................................................................................................................................................................................... 30
3.2. Luchtkwaliteit ........................................................................................................................................................................................... 30
3.3. HEPA-filter ............................................................................................................................................................................................... 30
3.4. Soorten luchtstroomkasten .................................................................................................................................................................... 30
3.5. Werken in een luchtstroomkast (LAF-kast) ........................................................................................................................................ 31
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, Celcultuur en immunochemie | [Author Name]
4. Microscopie .....................................................................................................................................................32
4.1. Geïnverteerde fase-contrast microscoop .......................................................................................................................................... 32
4.2. Epifluorescentie ....................................................................................................................................................................................... 32
4.3. GFP............................................................................................................................................................................................................ 32
5. Celteller......................................................................................................................................................32
Hoofdstuk 8: Sterilisatie en desinfectie ..................................................................................................................................... 33
1. Definities....................................................................................................................................................33
2. Gevoeligheid van kiemen tegen doding (!!) ...............................................................................................33
3. Steriliteit in industrie ..................................................................................................................................33
3.1. Farma-industrie ....................................................................................................................................................................................... 33
3.2. Onderzoekslabo’s .................................................................................................................................................................................. 33
4. Sterilisatieprocessen ..................................................................................................................................34
4.1. Stoomsterilisatie ...................................................................................................................................................................................... 34
4.2. Droge hitte sterilisatie ........................................................................................................................................................................... 36
4.3. Gassterilisatie ......................................................................................................................................................................................... 37
5. Straling ......................................................................................................................................................37
5.1. Niet-ioniserende straling = desinfectie ............................................................................................................................................. 37
5.2. Ioniserende straling ................................................................................................................................................................................ 38
6. Vloeistoffiltratie .........................................................................................................................................38
6.1. Soorten ..................................................................................................................................................................................................... 38
6.2. Materiaal ................................................................................................................................................................................................. 39
6.3. Nadelen filtratie ..................................................................................................................................................................................... 39
6.4. Wanneer gebruikt.................................................................................................................................................................................. 39
7. Validatie van sterilisatie ............................................................................................................................39
7.1. Fysische validatie ................................................................................................................................................................................... 39
7.2. Microbiologische validatie ................................................................................................................................................................... 39
7.3. Chemische indicatoren ........................................................................................................................................................................... 39
Hoofdstuk 9: Bioveiligheid .......................................................................................................................................................... 40
1. Inleiding.....................................................................................................................................................40
2. Risicoklassen .............................................................................................................................................40
3. Inperken van risico’s..................................................................................................................................40
3.1. Maatregelen............................................................................................................................................................................................ 40
3.2. Fysische inperking ................................................................................................................................................................................... 40
3.3. Werkvoorschriften .................................................................................................................................................................................. 40
Hoofdstuk 10: Contaminaties ..................................................................................................................................................... 41
1. Bronnen van contaminatie .........................................................................................................................41
1.1. De operator ............................................................................................................................................................................................. 41
1.2. De labo-omgeving ................................................................................................................................................................................. 41
1.3. Microbiologische veiligheidskast ......................................................................................................................................................... 41
1.4. Waterbad................................................................................................................................................................................................ 41
1.5. Incubatoren .............................................................................................................................................................................................. 41
1.6. Koude kamers ......................................................................................................................................................................................... 41
1.7. Steriel materiaal..................................................................................................................................................................................... 41
1.8. Gekregen cellijnen en weefsels .......................................................................................................................................................... 42
1.9. Vloeibare stikstof ................................................................................................................................................................................... 42
2. Typen contaminatie ...................................................................................................................................42
2.1. Kruiscontaminatie .................................................................................................................................................................................... 42
2.2. Contaminatie met micro-organismen .................................................................................................................................................. 42
Hoofdstuk 11: Karakterisering ................................................................................................................................................... 44
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