Tus Nqi Tsawg Tus Nqi Bioprinter: 13 Kauj Ruam (nrog Duab)

2024 Tus sau: John Day | [email protected]. Kawg hloov kho: 2024-01-30 09:26

Peb yog pab pawg tshawb fawb qib qis hauv UC Davis. Peb yog ib feem ntawm BioInnovation Group, uas ua haujlwm hauv TEAM Molecular Prototyping thiab BioInnovation Lab (Tus Kws Pab Tswv Yim Dr. Marc Facciotti, thiab Andrew Yao, MS). Lub chaw kuaj ntshav sib koom ua ke cov tub ntxhais kawm ntawm ntau haiv neeg sib txawv los ua haujlwm ntawm txoj haujlwm no (mech/chemical/biomed engineering).

Me ntsis keeb kwm yav dhau los ntawm txoj haujlwm no yog tias peb pib luam tawm cov noob nplej transgenic hauv kev sib koom tes nrog Dr. Karen McDonald ntawm ChemE chav haujlwm nrog lub hom phiaj txhawm rau txhim kho tus nqi qis bioprinter los ua bioprinting nkag mus tau yooj yim rau cov chaw tshawb fawb. Tam sim no, cov bioprinters qis kawg raug nqi kwv yees li $ 10, 000 thaum lub tshuab bioprinters qib siab raug nqi kwv yees li $ 170, 000. Qhov sib txawv, peb lub tshuab luam ntawv tuaj yeem tsim tau kwv yees li $ 375.

Khoom siv

Qhov:

1. Ramps 1.4:
2. Arduino mega 2560:
3. Stepper lub cev muaj zog tsav tsheb:
4. Stepper lub cev muaj zog ntxiv (xaiv tau)
5. Tus nqaj hlau 2 hauv X 1 hauv
6. Maker beam txuas cov cuab yeej
7. M3 screws ntau qhov sib txawv
8. M3 noob txiv x2
9. 8 hli threaded pas nrig
10. 8mm noob
11. 608 cev
12. Phau ntawv tuav
13. Filament
14. Monoprice V2
15. Zip kev sib txuas
16. M3 cua sov teeb txiv ntseej 2 hli dav

Cov Cuab Yeej:

1. Laum cov khoom ntawm ntau qhov ntau thiab tsawg
2. Tes Laum
3. Laum xovxwm
4. Hacksaw
5. Soldering hlau + solder
6. Hlau stripper
7. Koob qhov ntswg pliers
8. Hex tuav ntau qhov ntau thiab tsawg

Cov khoom siv lab:

1. Petri tais ~ 70mm txoj kab uas hla
2. 60 ml koob txhaj tshuaj nrog Luer-lock taub
3. 10 ml koob txhaj tshuaj nrog Luer-lock taub
4. Luer-xauv fittings
5. Tubing rau fittings
6. T Connector rau tubing
7. Centrifuge
8. Centrifuge raj 60ml
9. Nplai
10. Ntsuas lub nkoj
11. Autoclave
12. Beakers
13. Kawm tiav lub tog raj kheej
14. 0.1M CaCl2 daws
15. Agarose
16. Alginate
17. Methylcellulose
18. Sucrose

Software:

1. Fusion 360 lossis Solidworks
2. Arduino IDE Cov
3. Repetier tus tswv
4. Ultimaker Cura 4

Kauj Ruam 1: Xaiv Lub Tshuab Luam Ntawv 3D

Peb xaiv Monoprice MP Xaiv Mini 3D Tshuab Luam Ntawv V2 ua lub tshuab luam ntawv 3D pib. Lub tshuab luam ntawv no tau raug xaiv vim nws tus nqi qis thiab muaj ntau. Ib qho ntxiv, tus qauv 3D zoo heev ntawm lub tshuab luam ntawv twb muaj lawm uas ua rau tsim tau yooj yim dua. Cov lus qhia no yuav raug txiav rau lub tshuab luam ntawv tshwj xeeb no tab sis cov txheej txheem zoo sib xws tuaj yeem siv los hloov lwm yam FDM tshuab luam ntawv thiab CNC tshuab.

Tus qauv raug siab:

Kauj Ruam 2: Luam Ntawv 3D

Ua ntej kev tshem tawm ntawm Monoprice lub tshuab luam ntawv, ntau qhov yuav tsum tau luam tawm 3D rau kev hloov kho ntawm 3D tshuab luam ntawv. Muaj cov qauv ntawm cov tshuaj txau extruders, ib qho uas xav tau epoxy thiab ib qho uas tsis ua. Qhov uas xav tau epoxy yog qhov sib dua tab sis nyuaj rau sib sau ua ke.

Kauj Ruam 3: Npaj Tshuab Luam Ntawv rau Kev Hloov Kho

Lub vaj huam sib luag pem hauv ntej, npog hauv qab thiab lub vaj huam sib luag tswj yuav tsum raug tshem tawm. Thaum hauv qab tau raug tshem tawm, tshem tag nrho cov khoom siv hluav taws xob los ntawm pawg tswj hwm thiab tshem tawm pawg tswj hwm.

Kauj Ruam 4: Sib Hloov Mount

Lub Cev 1 thiab Lub Cev 14 txhua tus xav tau ob lub txiv ntoo ua kom sov. Lub cev 1 tau teeb tsa rau ntawm lub tshuab luam ntawv thav duab los ntawm ob lub M3 ntsia hlau zais hauv qab txoj siv. Cov ntsia liaj qhov rooj tuaj yeem tshwm sim los ntawm kev tshem txoj hlua nruj thiab rub txoj siv mus rau ib sab.

Kauj Ruam 5: Z Axis Hloov

Z-axis hloov tau hloov chaw kom txhua rab koob ntev tuaj yeem siv tau thaum lub sijhawm nyob ua ke yam tsis tau them nyiaj hauv software. Qhov hloov pauv yuav tsum tau teeb tsa nrog 2 M3 ntsia hlau rau lub tshuab luam ntawv chassis ncaj qha hauv qab lub taub hau luam ntawv kom ze rau ntawm lub txaj luam ntawv li sai tau.

Kauj Ruam 6: Txuas kab

Kev txuas hluav taws xob tau ua tiav raws li Ramps 1.4 cov qauv. Tsuas ua raws daim duab kab thaiv. Txiav tawm thiab xaim xov hlau raws li xav tau rau cov davhlau ya nyob twg. Qee cov xov hlau yuav tsum tau txuas ntxiv.

Kauj Ruam 7: Epoxy Extruder

Thaum lub tshuab extruder no siv sijhawm tsawg dua los luam tawm, nws siv epoxy uas ua rau lub sijhawm tsim tag nrho mus txog 24 teev. Lub 8mm threaded rod yuav tsum tau epoxied rau 608 cov kabmob thiab cov kabmob yuav tsum yog epoxied rau 3D luam tawm daim Lub cev 21. Tsis tas li ntawd, txiv ntseej rau cov xov threaded yuav tsum tau epoxied rau Lub Cev 40. Thaum epoxy tau kho tas lawm, roj hmab cov lus qhia los ntawm 60ml thiab 10 ml koob txhaj tshuaj tuaj yeem haum rau Lub Cev 9 thiab Lub Cev 21, feem. Qhov tsim nyog T haum tsis tuaj yeem pom yog li cov khoom nyoos tau tsim los ntawm 6mm tooj dag tubing thiab cov laug. Lub tshuab extruder ua raws li cov txheej txheem hydraulic uas thawb Bioink tawm ntawm chav qis ntawm 10 ml koob txhaj tshuaj. Huab cua tuaj yeem khiav tawm ntawm lub kaw lus los ntawm kev ua kom co lub raj thaum tuav T haum ntawm qhov chaw siab tshaj.

Kauj Ruam 8: Tsis Siv Neeg Muab Tshuaj Txhuam Extruder

Lub tshuab extruder no tuaj yeem yooj yim raug ntsia ua ke. Qhov tsis zoo ntawm cov extruder no yog tias nws yog qhov hnyav thiab muaj qhov siab rov qab.

Kauj Ruam 9: Kauj Ruam 9: Arduino Firmware

Arduino xav tau firmware los khiav cov tsav tsheb thiab lwm yam khoom siv hluav taws xob. Peb xaiv Marlin raws li nws yog pub dawb, hloov kho yooj yim nrog Arduino IDE thiab txhawb tau zoo. Peb tau hloov kho lub khoos phis tawj rau peb cov cuab yeej tshwj xeeb tab sis nws yooj yim heev rau hloov pauv rau lwm lub tshuab luam ntawv vim tias txhua tus lej tau hais tawm thiab piav qhia meej. Ob npaug nias rau MonopriceV2BioprinterFirmware.ino cov ntaub ntawv los qhib cov ntaub ntawv teeb tsa marlin.

Kauj ruam 10: Cura Profile

Cov ntaub ntawv Cura tuaj yeem raug xa mus rau Ultimaker Cura 4.0.0 thiab siv los ua kom thaj tsam thaj tsam siab rau siv rau hauv cov tshuaj tua kab mob. Cov cim ntawm Gcode rau lub tshuab luam ntawv tseem muaj kev sim siab thiab xav tau kev ua siab ntev ntau. Kuj tseem txuas nrog yog qhov ntsuas gcode rau lub tshuab ua kom muaj zog ntau ntxiv.

Kauj ruam 11: Hloov Pib G-code

Muab tus lej no tso rau hauv pib G-code teeb tsa:

G1 Z 15

G28

G1 Z20 F3000

G92 Z33.7

G90

M82

G92e0 ua

Hauv Repetier, txhawm rau hloov kho pib Gcode mus rau lub tshuab txiav-> Kho Kom Zoo Nkauj-> G-lej-> pib G-codes. Nws yuav tsum tau hloov kho G92 Z tus nqi rau txhua rooj plaub tshwj xeeb. Maj mam nce tus nqi kom txog thaum rab koob xav tau qhov deb ntawm Petri phaj nto thaum pib luam ntawv.

Kauj Ruam 12: Ua Bioink

Cov txheej txheem txhawm rau txhim kho Bioink haum rau daim ntawv thov yog nyuaj. Nov yog txheej txheem uas peb tau ua raws:

Ntsiab lus

Lub hydrogel tsim nyog rau shear-sensitive cog hlwb thiab muaj qhib macropores kom tso cai rau kis. Hydrogel yog tsim los ntawm kev ua kom yaj agarose, alginate, methylcellulose, thiab sucrose hauv dej deionized thiab ntxiv cov cell. Cov gel yog khov kom txog thaum nws tau kho nrog 0.1M calcium chloride, uas ua rau nws khov kho. Cov tshuaj calcium chloride tua kab hla kev sib txuas nrog alginate kom nws ruaj khov. Alginate yog lub hauv paus ntawm cov gel, methylcellulose homogenizes cov gel, thiab agarose muab ntau tus qauv txij li nws cov gel ntawm chav sov. Cov sucrose muab zaub mov rau cov cell txuas ntxiv mus ntxiv hauv cov hydrogel.

Cov lus piav qhia luv luv ntawm qee qhov kev sim los txheeb xyuas cov gel

Peb tau sim cov hydrogels sib txawv nrog ntau qhov sib txawv ntawm agarose thiab sau nws qhov sib xws, nws yooj yim npaum li cas nws luam tawm, thiab seb nws puas los yog ntab hauv cov tshuaj tua kab. Kev txo qis alginate feem pua ua rau cov kua ua kua ntau dhau thiab nws tsis muaj peev xwm khaws nws cov duab tom qab luam ntawv. Ua kom cov alginate feem pua ua rau cov tshuaj tua kab ua haujlwm sai sai, tias cov gel yuav kho ua ntej lo rau saum txheej. Hydrogel uas tuav nws lub cev thiab tsis kho sai heev tau tsim los siv 2.8 wt% alginate.

Yuav ua li cas txhawm rau txhim kho hydrogel

Khoom siv

Agarose (0.9 wt %)

Alginate (2.8 wt %)

Methylcellulose (3.0 wt%)

Sucrose (3.0 wt%)

Calcium Chloride.1M (147.001 g/mol)

ddH20

cell sib sau ua ke

2 Ntxuav & Qhuav Beakers

1 Sib tov Spatula

Txhuas Ntawv Nyiaj

Yas Luj Ntawv

Kawm tiav Lub tog raj kheej

Txheej txheem

Ua Hydrogel:

1. Ntsuas tawm qhov tshwj xeeb ntawm ddH20 raws li ntau npaum li cas cov kua tshuaj uas koj xav npaj. Siv lub tog raj kheej kawm tiav kom tau txais qhov ntim tshwj xeeb ntawm ddH20.
2. Cov tshuaj hydrogel yuav muaj Alginate (2.8 wt %)), Agarose (0.9 wt %), sucrose (3 wt %), thiab methylcellulose (3 wt %). Ib feem ntawm cov khoom ntawm cov tshuaj hydrogel yuav raug ntsuas siv cov yas hnyav ntawv.
3. Thaum ua tiav qhov hnyav tag nrho cov khoom, ntxiv ddh20, sucrose, agarose, thiab kawg sodium alginate rau ib qho ntawm cov dej khov qhuav. Swirl los sib tov tab sis tsis txhob siv lub spatula los sib tov vim tias cov hmoov yuav lo rau ntawm cov spatula.
4. Thaum sib xyaw, qhwv sab saum toj ntawm lub beaker nrog aluminium ntawv ci kom raug thiab sau lub beaker. Ntxiv ib daim kab xev autoclave rau saum cov ntawv ci.
5. Muab cov methylcellulose seem tso rau hauv lwm lub tais qhuav thiab qhwv nws hauv txhuas ntawv ci zoo li lub khob yas dhau los. Sau daim ntawv no beaker thiab ntxiv ib daim kab xev autoclave rau saum cov ntawv ci.
6. Qhwv 1 spatula hauv ntawv ci txhuas thiab xyuas kom tsis muaj leej twg pom. Ntxiv daim kab xev autoclave rau lub qhwv spatula.
7. Autoclave 2 lub raj mis thiab 1 rab diav ntawm 121 C rau 20 feeb thaum lub sijhawm ua kom tsis muaj menyuam. Tsis txhob siv AUTOCLAVE hauv STERILE & DRY CYCLE.
8. Thaum lub voj voos autoclave ua tiav, cia cov gel txias mus rau qhov kub hauv chav thiab thaum nws tau mus txog nws, pib ua haujlwm hauv Txhab Kev Nyab Xeeb Kev Nyab Xeeb.
9. Nco ntsoov ntxuav koj txhais tes thiab txhais caj npab thiab siv cov txheej txheem aseptic kom raug thaum ua haujlwm hauv lub txee rau kev nyab xeeb. Kuj TSWV YIM kom tsis txhob cuam tshuam ncaj qha nrog cov khoom uas yuav kov cov gel lossis ze rau cov gel (ex: sib tov kawg ntawm spatula, lossis thaj tsam ntawm aluminium foils uas zaum saum cov gel)
10. Hauv lub thawv ntim khoom noj kom nyab xeeb sib tov methylcellulose rau hauv cov gel kom tau txais homogeneous kis. Thaum ua tiav kev sib tov, rov sau rau sab saum toj ntawm cov tshuaj sib tov thiab tso rau hauv lub tub yees thaum hmo ntuj.
11. Los ntawm no cov gel tuaj yeem siv rau kev qhia ntawm lub cell lossis rau lwm yam siv xws li luam ntawv.

Ntxiv Cells:

1. Lim cov cell kom lawv zoo ib yam. Peb cov txheej txheem rau kev lim dej yog

   Maj mam txhuam cov cell tawm ntawm lub tais petri thiab siv 380 micrometer sab cib los lim cov cell.

2. Maj mam muab cov cell lim dej hauv cov tshuaj hydrogel siv lub taub hau ncaj kom tsis txhob poob qhov sib xyaw (uas tau ua autoclaved).
3. Tom qab sib xyaw cov cell centrifuge tawm ua npuas
4. Los ntawm no hydrogel ua tiav thiab tuaj yeem siv rau kev luam ntawv, kho, thiab kev sim yav tom ntej.

Yuav ua li cas txhawm rau txhim kho cov tshuaj tua kab (0.1M Calcium chloride, CaCl2)

Khoom siv

Calcium chloride

ddH20

Sucrose (3 feem pua %)

Cov txheej txheem (ua kom 1L tua tshuaj)

1. Ntsuas 147.01g calcium chloride, 30mL sucrose, thiab 1L ddH20.
2. Sib tov cov tshuaj calcium chloride, sucrose, thiab ddH20 hauv lub tais loj lossis ntim khoom.
3. Muab cov gel tso rau hauv cov tshuaj tua kom ntev li 10 feeb kom kho tau.

Kauj ruam 13: Luam tawm

Hauv kev xav, Bioprinting yog qhov yooj yim heev; txawm li cas los xij, hauv kev xyaum, muaj ntau yam uas tuaj yeem ua rau tsis ua tiav. Nrog cov gel no, peb tau pom tias ntau yam tuaj yeem ua tiav kom ua tiav zoo tshaj plaws rau peb daim ntawv thov:

1. Siv me me ntawm CaCl2 daws kom ib nrab kho cov gel thaum luam ntawv,
2. Siv daim ntawv so tes hauv qab ntawm lub tais petri txhawm rau txhawm rau ua kom khov
3. Siv daim ntawv so tes kom sib npaug me me ntawm CaCl2 hla tag nrho cov ntawv luam tawm
4. siv flowrate slider hauv Repetier txhawm rau nrhiav qhov tseeb ntws

Rau cov ntawv thov sib txawv thiab sib txawv gels, cov txheej txheem sib txawv yuav tsum tau siv. Peb cov txheej txheem tau tsim nyob rau ntau lub hlis. Kev ua siab ntev yog qhov tseem ceeb.

Hmoov zoo yog tias koj sim ua txoj haujlwm no thiab xav nug txhua yam lus nug.

Thawj qhov khoom plig hauv Arduino Contest 2019