dynamic BIOSENSORS heliX cyto Sistemụ nyocha nyocha ụlọ nyocha zuru oke

Nkọwapụta

• Aha ngwaahịa: heliX cyto
• Onye nrụpụta: Dynamic Biosensors GmbH
• Ụdị: 1.0

Ozi ngwaahịa

• Emebere sistemu cyto heliX maka otu cell Interaction Cytometry (scIC) iji tụọ njikọ njikọ nke ụmụ irighiri ihe akpọrọ fluorescent iji lekwasị anya n'elu cell n'ụzọ edoziri oge.

Ntuziaka ojiji ngwaahịa

Iji heliXcyto Chip

• Ihe mgbawa heliXcyto nwere otu ọwa microfluidic nwere oghere electrode maka ijide cell na nha. Gbaa mbọ hụ na etinyere mgbawa ahụ nke ọma wee soro ntuziaka maka ọnyà sel.

Chip mmezi

• Jiri Chip Mmezi maka ihicha, nnwale, na ọrụ mmezi naanị. Zere iji ya maka nnwale iji gbochie mmetọ.

Ihe nchekwa na-agba ọsọ

• Jiri ihe nchekwa na-agba ọsọ 1 (RB 1) n'oge nha. Rụtụ aka na mpempe akwụkwọ ndakọrịta scIC na ntuziaka bụ isi maka ozi zuru ezu.

Nyocha Sensorgram

• Lelee sensọgram ozugbo na ngwa heliOS n'oge nha. Nyochaa nzaghachi fluorescent ka oge na-aga wepụta ọnụego kinetic.

Mmasị

• Tụlee iji passivation dị ka nzọụkwụ nhọrọ iji gbochie mgbasa ozi nyocha site na ịmalite reagent passivation na mgbawa.

Normalization

• Normalization bụ usoro iji hazie data maka ntụnyere na nyocha. Soro usoro nhazigharị dịka ntuziaka onye ọrụ si dị.

Okwu mmalite

• Ntuziaka a pụtara dị ka nkenkeview nke usoro heliXcyto na ike ya. A na-agbasawanye isiakwụkwọ ndị a na isi akwụkwọ ntuziaka heliXcyto achọtara na https://www.dynamic-biosensors.com/helios-download-latest/

heliXcyto Terminology otu cell mmekọrịta Cytometry

• Otu cell Interaction Cytometry (scIC) bụ nkà na ụzụ na-atụta njikọ njikọ nke molekul akpọrọ fluorescently (analyte) na ebumnuche (ligand) n'elu cell site na ịdekọ akara fluorescent n'oge edoziri.

Nyochaa

• Analyte bụ molekul akpọrọ fluorescent na usoro mkpanaka nke nwere ike jikọta na ebumnuche dị n'elu cell.

Ligand

• Ligand” bụ aha e ji na ngwanrọ heliOS kọwaa ebumnuche dị n'elu sel nke onye nyocha nwere ike ijikọ ya.

Ọnyà cell

• Ọnyà cell bụ ihe na-eruba-na-asọba, oghere polymer dakọtara na mgbawa heliXcyto. Emebere ha iji weghara ma mebie sel ndị nwere nha dị iche site na ihe dịka 6 ruo 25 µm na ọwa eruba. Ọnyà cell dị na nha atọ dị iche iche: obere, ọkara na nnukwu.

heliXcyto Chip

• Chips heliXcyto nwere otu ọwa microfluidic nwere oghere n'akụkụ nke ọ bụla. Ebe abụọ electrode ọla edo dị n'etiti ọwa.
• Otu ntụpọ na-eje ozi dị ka ntụpọ ntụaka, ebe nke abụọ na-eburu 3D biocompatible polymer cages maka ijide cell ma na-eje ozi dị ka ntụpọ ntụ. Ebe nha nwere ike ịnwe ọnyà 1 ma ọ bụ 5 nke otu ụdị.
• Ebe ntụnyere aka na-enye ohere ntutụ aka ozugbo nke mgbama fluorescent anakọtara. Ihe mgbawa heliXcyto bụ nke enwere ike iji ya mee ihe ma tụfuo ya.

Chip mmezi

• Chip Mmezi bụ otu mgbawa microfluidic ọwa ejiri maka nhicha, nnwale na arụmọrụ niile.
• Arụmọrụ ndị a gụnyere usoro dị ọcha & ihi ụra, teta n'ụra na usoro ihe omume, ịsacha sistem na ule Fluidics. E kwesịghị iji mgbawa a mee nnwale n'ezie na ngwọta cell/protein iji gbochie mmetọ ọzọ nke mgbawa.

Ihe nchekwa na-agba ọsọ

• Ihe nchekwa na-agba ọsọ bụ ihe nchekwa ejiri na heliXcyto n'oge nha. A na-akwadokarị iji ihe nchekwa na-agba ọsọ 1 (RB 1).
• Biko rụtụ aka na mpempe akwụkwọ ndakọrịta scIC achọtara na ntuziaka bụ isi maka ozi ndị ọzọ.

Sensorgram

• Ihe mmetụta sensọ scIC bụ atụmatụ nzaghachi fluorescence na ọnụọgụ kwa sekọnd (cps) ka oge na-aga, na-egosi ọganihu nke mmekọrịta na ma ọ bụ na sel. Usoro a nwere ike ịbụ viewed na ozugbo na heliOS n'oge nha.
• Mgbe ị na-enyocha ihe mmetụta sensọ, a na-ekpebi nkọwa kacha mma maka akara fluorescent a na-ahụ anya ma wepụta ọnụego kinetic (kon, koff).

Mmasị

• Passivation bụ nzọụkwụ nhọrọ na-agba ọsọ, ebe a na-agbanye ihe reagent passivation n'ime mgbawa ma tinye ya ka ọ gbochie elu. Nke a nwere ike inye aka igbochi mgbasa ozi nke nyocha na ihe mgbawa.

Normalization

• A na-eji usoro ngbanwe nke ọma were were nweta ntakịrị ọdịiche dị na mgbaama n'ihi nchịkọta mgbaama sitere na ebe nha ọ bụla site na onye nchọta dị iche. A na-agbanye ihe na-acha ọkụ fluorescent nke mkpokọta akọwapụtara (dabere na nchịkọta nyocha kachasị elu na ogo nyocha nyocha) na mmalite nke nyocha. A na-etinye usoro ngbanwe a na-akpaghị aka na usoro nha, yana a na-eji ọnụ ọgụgụ kachasị elu nke normalize na-emezi ozugbo maka ndịiche dị n'etiti ihe nchọpụta n'oge nyocha data, tupu ntugharị oge.

Ụkpụrụ nha nke scIC

Ngwa scIC

• Otu cell Interaction Cytometry (scIC) na-atụ ọnụego kinetic na njikọ nke nyocha nke akpọrọ fluorescent na-ejikọta ma na-ewepụ ya na ebumnuche ya ozugbo na sel dị ndụ.
• Nchọpụta nke nyocha na scIC bụ nke na-adabere na ya, ya mere ọ dị mkpa na molecule ọ bụla sitere na sub-nm ruo> 100 nm ma dị mkpa na klas molecule niile sitere na obere ụmụ irighiri, peptides, aptamers, nanobodies, affibodies, antibodies, bispecific antibody formats, proteins, proteins, proteins, protein, protein, protein, protein, protein, protein, protein, protein, protein, protein, Lixosomes. nanoparticles, na nje virus.

Teknụzụ scIC nwere ike ilebara mpaghara ngwa ndị a:

• nha nke mgbaàmà amplitudes na enyo enyo
• Kinetic analysis of kon na koff ọnụego
• affinity na avidity nwale site koff na biphasic dabara ụdị
• Specificity ule
• ikwu quantification nke akpụkpọ ahụ protein n'elu cell
• epitope binning na asọmpi ọmụmụ
• ihe mgbochi mgbochi
• nyocha nke internalization nke protein ezubere iche

ike heliXcyto Hardware

Sistemụ Fluidic

• A na-enye usoro mmiri mmiri nke heliXcyto site na karama ihe nchekwa 3 dị iche iche, na-eme ka mgbanwe dị iche iche na-agba ọsọ na nchekwa nchekwa. Enwere ike ịtọ nfuli dị na heliXcyto ka ọ na-erute ọnụego n'etiti 20 na 500 µL/min, na-eri nri dị iche iche s.ample na assay chọrọ. Ọwa mmiri na-agbawa nke mgbawa na-ejikọta na usoro mmiri mmiri site na oghere abụọ. Ejikọtara oghere n'akụkụ aka ekpe na sample, ihe nchekwa, na ahịrị ịsacha, ebe oghere akụkụ aka nri jikọtara ya na ahịrị mmeghari.
• Usoro mmiri nke ụzọ abụọ a na-enyere aka ijide cell kwụsiri ike ka ọ na-agba ọsọ dị iche iche nke nha ma mechaa nweghachi mgbawa maka mgbawa in-assay reusability.
• Onyonyo 1. Ngwakọta mgbawa na sistemu Fluidic

Sistemụ ngwa anya

• Ihe mgbaàmà fluorescence na-enwe obi ụtọ site na diodes na-acha uhie uhie na akwụkwọ ndụ akwụkwọ ndụ (LED) wee chọpụta ya site na ọnụ ọgụgụ foto anọ, nke na-anakọta akara uhie na akwụkwọ ndụ akwụkwọ ndụ site na ebe ọ bụla n'ime omimi zuru oke nke ọwa.
• Enwere ike nyochaa akara abụọ nọọrọ onwe ha n'otu oge n'otu ebe nha, na-enye ohere ka enwere ike tụọ mmekọrịta abụọ n'otu oge na nhazi nyocha nke agba abụọ.
• Onyonyo 2. Nhazi ngwa anya
• A na-ejikwa ihe nchọta dị iche iche na-ahụ maka mkpokọta mgbaama sitere na ebe nha ọ bụla, nke nwere ike bute ntakịrị ọdịiche dị na ọnụ ọgụgụ raw. Iji zaa ajụjụ maka nke a, a na-etinye usoro nhazigharị anya na-akpaghị aka na nyocha ndị na-emepụta data.
• Na mgbakwunye na ọnụ ọgụgụ otu foto, akụrụngwa a nwere igwefoto CCD na microscope ìhè na-egosipụta. Ngwá ọrụ ndị a na-enye ohere maka onyonyo cell ozugbo, na-enyere aka nyochaa iguzosi ike n'ezi ihe cell na ịdị mma nyocha n'oge nha.
• Nhazi a jikọtara ọnụ na-eme ka a na-enyocha ma mmekọrịta na ọnọdụ ndu, na-enye nyocha zuru oke nke nnwale ahụ.

SIC usoro ọrụ

Enwere ike imebi usoro nha scIC n'ime usoro isi atọ

1. Nleba anya na heliOS: Ntụle ọ bụla na-amalite site n'ichepụta nnwale na ngwa heliOS. A na-ahazi usoro nyocha nke nyocha site n'ịhọrọ ụzọ ndị eburu ụzọ na-emezi ihe nyocha, dị ka ahịrị cell ejiri, ntinye nyocha, na ọnọdụ nchekwa. heliOS na-akpaghị aka na-agbakọ ọnụ ọgụgụ nke mkpụrụ ndụ, nyocha, ihe nchekwa na ihe ngwọta ndị ọzọ dị mkpa maka nha, na-eme ka usoro nkwadebe ahụ dịkwuo mma.
2. Nkwadebe nke nchekwa, nyocha na sel: Mgbe emechara ihe nnwale ahụ, a na-akwado ihe ndị achọrọ dịka nkọwapụta nke heliOS nyere. A na-ebunye ihe nchekwa, ndị nyocha na sel n'ime akpaaka ngwaampler.
3. Ntụle na nyocha data: Sistemu ahụ na-arụ usoro akwadoro nke nha mmekọrịta ozugbo na-achọghị enyemaka ọzọ n'aka onye ọrụ. Enwere ike nyochaa data mgbe nha ka na-agba ọsọ iji nweta nghọta ozugbo na nsonaazụ ya.

Nyocha scIC na heliOS

Na mgbakwunye na nyocha nke na-emepụta data, ngwanro ahụ na-enyekwa ụzọ maka ịnwale mgbawa, nhicha na nhazi ngwa.

Tebụl 1. Nnwale na-emepụta data enwetara

SIC Kinetics	Na-atụkwa kinetics zuru oke na sel nwere njikọ nyocha nwere ike ịhazigharị n'etiti mkpokọta 1 na 5, na-esochi otu nkewa mgbe itinye uche kacha elu. Ma ọwa akwụkwọ ndụ akwụkwọ ndụ ma ọ bụ ọbara ọbara. Nwere nhọrọ maka naanị nyocha nyocha.
SIC Kinetics - Agba abụọ	Na-atụkwa kinetics zuru oke na sel nwere njikọ nyocha nwere ike ịhazigharị n'etiti mkpokọta 1 na 5, na-esochi otu nkewa mgbe itinye uche kacha elu. Agbanyere ọwa akwụkwọ ndụ akwụkwọ ndụ na uhie n'otu oge. Nwere nhọrọ maka naanị nyocha nyocha.
Nkewa nke scIC - Agba abụọ	Na-atụ nkewa nke onye nyocha site na sel ndị ejirila nyocha nke akpọrọ fluorescent nwere ọwa akwụkwọ ndụ akwụkwọ ndụ na/ma ọ bụ uhie.

Nkwadebe na mmepe nyocha maka nha scIC

Enwere isi ihe ole na ole ị ga-atụle tupu ịtọọ nha scIC

• Nleba anya nyocha, Degree of Labeling, Properties, and quality.
• Ngwọta Normalisation na ike mkpali.
• Ogo cell na njikwa.

Ntụle nyocha na ogo nke ịde aha

• Dị ka usoro nha nha nha nha anya, anyị na-akwado ịhọrọ oke mkpokọta mkpokọta nyocha nke 0.1x ruo 10x nke a na-atụ anya nha nhata mgbe niile (Kd) maka ọtụtụ kinetics itinye uche. Maka otu kinetics itinye uche ma ọ bụ nha nkewa, ntinye nyocha kwesịrị ịdị n'etiti 1x na 10x Kd a na-atụ anya ya. A na-agbakọ olu nyocha achọrọ maka oge mkpakọrịta nkeji iri na ọsọ ọsọ nke 25 µL/min na-agbakọta ihe dịka 300 µL.
• Degree of Labeling (DOL) maka onye nyocha kwesịrị ịbụ 1, n'agbanyeghị ụkpụrụ DOL nke 2 ma ọ bụ 3 ka anabatara. Otú ọ dị, mara na ọnụ ọgụgụ dị elu nke akara akara na onye nyocha nwere ike imetụta njirimara njide ya, ma nwee ike ịbawanye ohere nke nchịkọta ma ọ bụ nke a na-akọwaghị. Iji nweta ezigbo DOL, soro ụkpụrụ enyere na ngwa Labeling nwere ihe na-acha uhie uhie ma ọ bụ akwụkwọ ndụ akwụkwọ ndụ sitere na ahịrị heliXcyto reagents.
  Normalization
• Nkịtị bụ nzọụkwụ mbụ n'ime usoro mmepụta data niile. Ọ na-akwụ ụgwọ maka ntakịrị mgbanwe mgbaama kpatara site na iji ihe nchọpụta dị iche iche maka ebe nha ọ bụla. N'ime usoro a, a na-agbanye ihe na-acha fluorescent na ntinye aka nke onye ọrụ na mmalite nke nyocha ahụ.
• A na-agbakọ mkpokọta Ngwọta Normalisation site n'ịba ụba mkpokọta nyocha nke akpọrọ fluorescent nke ejiri na nha site na Degree's Degree of Labeling (DOL). Nke a itinye uche na-eduzi LED excitation ike etinyere n'oge nha. Ebumnobi bụ maka Normalisation Ngwọta ọnụ ọgụgụ kasị elu iru ihe dị ka otu mgbama fluorescence amplitude dị ka mkpokọta nyocha kachasị elu.
• Rụtụ aka na eserese ịta ahụhụ Normalisation Solution na ntuziaka bụ isi ka ịtọ ụkpụrụ mbụ. Rịba ama na eserese ahụ na-eduzi ntọala mbụ, mana enwere ike gbanwee ike mkpali ọkụ ọkụ n'oge mmepe nyocha.

Ogo Cell na Nkwadebe

• Ịdị ndụ cell kwesịrị ịdị n'elu 95%, na mkpụrụ ndụ kwesịrị ịdị n'ọnọdụ dị mma. Maka sel ndị na-esote, anyị na-akwado ịrụ ọrụ na sel ndị nwere 50-70% confluent.
• Maka mkpụrụ ndụ nkwusioru, anyị na-akwado iwe ihe ubi n'oge etiti oge nke uto. 50 µl nke nkwusioru cell na 1E + 06 sel/mL dị mkpa kwa ọsọ.

Ngwunye mkpụrụ ndụ nkwusioru

1. Centrifuge ihe ruru. 2E + 06 cell na 300 g maka nkeji 7 iji wepụ mgbasa ozi omenala cell. Tụfuo ihe dị elu.
2. Sachaa mkpụrụ ndụ otu ugboro site na ịmaliteghachi pellet cell n'ihe dị ka. 1 ml nke DPBS. Centrifuge nkwusioru ahụ na 400 g maka nkeji 5 ka pellet sel ndị dị ndụ. Jiri nlezianya tụfuo ihe dị elu iji wepụ iberibe na mgbasa ozi.
3. Jiri nwayọ kwughachi na 1 ml nke DPBS, bufee nkwusioru n'elu strainer cell wee mebie nkwusioru ahụ site na ike ndọda.
4. Tụọ mkpokọta cell nke nkwusioru a kpasuru iwe wee gbanwee gaa na 1E+06 sel/mL.
   • NDỤMỌDỤ: Ụfọdụ mkpụrụ ndụ nkwusioru na-etolite ụyọkọ. Jiri nwayọ kwughachi ụyọkọ ma gụnye usoro mgbakasị ahụ tupu nzọụkwụ mbụ centrifugation.
   • Jiri ndụmọdụ pipette obosara obosara mgbe ị na-ejikwa nkwusioru cell ka ịzenarị ike dị elu n'oge mbufe ma ọ bụ nkwụghachi.

Na-ewe ihe ubi sel agbakwunyere

1. Jiri DPBS sachaa mkpụrụ ndụ.
2. Kewapụ mkpụrụ ndụ na flask cultureing site na iji reagent dissociation masịrị gị (dịka TrypLE).
3. Weghachite mkpụrụ ndụ kewapụrụ na olu mgbasa ozi masịrị wee jiri strainer cell 30 nm yochaa.
4. Nhọrọ, tinye EDTA ma ọ bụrụ na sel ndị na-akwadosi ike (ntụkwasị uche ikpeazụ 5 mM).
5. Centrifuge mkpụrụ ndụ na 300 g maka nkeji 7 iji wepụ mgbasa ozi omenala cell. Tụfuo ihe dị elu.
6. Weghachite mkpụrụ ndụ na 1 ml nke DPBS.
7. Tụọ mkpokọta mkpụrụ ndụ wee gbanwee ka 1E+06 sel/mL.
   • NDỤMỌDỤ: Enwere ike iji mgbasa ozi cell gbazere 1:4 na DPBS iji tinye nri na cell samples n'ihe banyere sel nwere mmetụta ma ọ bụ ogologo ọrụ nyocha ọrụ.

Ndozi cell

1. Centrifuge ruo sel 10E+06 na 300 g maka nkeji 7 iji wepụ mgbasa ozi omenala cell. Tụfuo ihe dị elu.
2. Weghachite pellet cell na 1 mL PBS, centrifuge ma tụfuo ihe dị elu.
3. Weghachite pellet cell na 500 µL PBS/2% PFA (62.5 µL nke 16% ngwọta PFA + 437.5 µL PBS).
4. Tinye mkpụrụ ndụ n'ime ụlọ maka nkeji iri (na-eji nwayọọ nwayọọ na-ama jijiji).
5. Tinye 500 µL nke PBS na centrifuge nkwusioru na 400 g maka nkeji 5 iji wepụ PFA.
6. Tụfuo ihe dị elu.
7. Weghachite mkpụrụ ndụ na 1 mL PBS, yochaa site na strainer cell 30 µm, centrifuge (400 g, 5 min) wee tụfuo ike.
8. Weghachite mkpụrụ ndụ ahụ n'ihe dịka 1 ml nke PBS (nhọrọ: gbanwee na ntinye ikpeazụ nke 5E+06 sel/mL).
9. Chekwaa sel edobere na 2-8 Celsius, jiri n'ime otu ọnwa.
   • Ọ bụrụ na ekwesịrị idozikwu sel, biko gbalite ego niile ka ọ dabara.
   • IHE: Enwere ike gbanwee ọsọ centrifugation n'oge owuwe ihe ubi nke mkpụrụ ndụ ka ọ bụrụ ụdị sel, ma ọ bụrụ na mkpụrụ ndụ nwere mmetụta ma na-ejikarị g-force.
   • Ntinye uche PFA nwere ike ịdị iche n'etiti 0.5 na 4%.

Ntọala nyocha nke scIC

A ga-etinye nyocha scIC ndị a na mmepe nyocha yana enwere ike ịmegharị ya n'ime usoro ọrụ nyocha ma emechaa.

A na-ekewa usoro mmepe nke assay n'ime usoro atọ dị iche iche:

1. Nnwale mgbawa cyto
2. Nyocha naanị ule
3. Nyocha nke Kinetics

Nnwale Cyto Chip

• Ọ dị mkpa ka enyocha ogo mgbawa ọhụrụ tupu iji mgbawa cyto na nha. Malite site n'ịgba ule Cyto Chip iche iche. Hụ Helix cyto mgbawa isi na ntuziaka bụ isi maka otu esi ahazi na nyochaa ule mgbawa.

Nyocha naanị ule

• Mmepe nke nyocha scIC na-amalite site na nyocha naanị nyocha, nke na-enyocha omume nke nyocha ndị akpọrọ fluorescently na elu mgbawa ọhụrụ na-enweghị mkpụrụ ndụ.
• Enwere ike ịmegharị ule a ugboro ugboro iji chọpụta nkwụsi ike nke ndị nyocha akara. Enwere ike ịhazi naanị ule nyocha nke scIC na nyocha Kinetics site n'ịkwanye igbe nyocha naanị n'okpuru ntọala cell.
• Maka ntule ogo nyocha, ntinye uche kachasị elu ejiri zuru oke ma nwee ike ịhazi ya site na iwepu ihe niile ma ewezuga otu n'ime mkpakọrịta dị na nyocha ahụ.

Kinetics Assay nhazi usoro ọrụ

• A na-ejikarị nyocha nke kinetics eme ihe n'ime usoro ọrụ akpaaka nke nwere ma nyocha na-emepụta data yana ihe mmezi. Ozugbo nyocha ahụ gafere ule njikwa mma Analyte naanị ule, enwere ike iji ya na nha na sel.
• To quantify kinetic rates reliably, the signal response during association should be concentration-dependent, increasing with higher analyte concentrations.
• Nleta kachasị elu kwesịrị iru ala dị larịị, nke na-egosi na enwetala njide kwụ ọtọ. Mkpakọrịta ahụ kwesịrị ịgba ọsọ ogologo oge iji ihichapụ akụkụ dị mkpa nke bound analyte (karịa 5%) iji mee ka usoro ahụ dabara adaba nke ọma.
• Jiri paramita ndabara na ọnọdụ nyocha dị ka mmalite ma gbanwee n'ihu dabere na nsonaazụ gị.

Example 1. akwadoro nyocha ọrụ usoro

Usoro nyocha nke nyocha na-enye ndị ọrụ ohere ịtụ nha na usoro mmezi dabere na mkpa nnwale ha akọwapụtara.

Usoro ọrụ nyocha nwere ike ịgụnye usoro ndị a n'usoro egosiri:

1. Isi (mgbawa mmezi)
2. SIC Kinetics (cells A, Analyte A, Cyto mgbawa)
3. Ncha Sistemu (mgbawa mmezi)
4. SIC Kinetics (cells B, Analyte A, Cyto mgbawa)
5. Ncha Sistemu (mgbawa mmezi)
6. Nyocha naanị scIC Kinetics ahaziri (Analyte A, Cyto mgbawa)
   • A na-eme usoro ịsa ahụ Prime na Sistemu na mgbawa Mmezi. A na-agụnye ịsacha Sistemu mgbe ị na-atụgharị gaa na ahịrị cell dị iche na tupu ịme nyocha naanị nyocha na njedebe nke ọrụ ahụ.
   • Maka nkọwa ndị ọzọ, rụtụ aka na ngalaba nke ịsacha sistemu Cyto na ntuziaka isi.
   • Mgbe ị na-akwụ ụgwọ nyocha nke kinetics na sel, tụlee ịdị ndụ cell, nke dabere na ahịrị sel.
   • Maka ahịrị cell nwere mmetụta, a na-atụ aro ka ị na-agba ọsọ naanị 1-2 nyocha kwa usoro ọrụ nyocha. Maka ahịrị cell ndị ọzọ na-agbanwe agbanwe, enwere ike ịdeba nyocha ọzọ.
   • IHE: Maka nyocha ndị ọzọ, kwadebe ngwọta cell na vials dị iche iche site n'ịkpọ mkpụrụ ndụ n'ụdị dị iche iche.
   • Gbaa mbọ hụ na reagents niile dị ọhụrụ ma kpochaa site na nzacha 0.22µm. N'ikpeazụ, tinye reagents akwadoro n'ime ngwá ọrụ dị ka heliOS gwara ya.
   • Enwere ike ịmalite ịgba ọsọ ahụ site na ịpị akara ngosi na-acha anụnụ anụnụ "Run" ma soro usoro ndị dị na Assay Start Wizard.
   • Ozugbo ịgba ọsọ ahụ malitere, ngwaọrụ ahụ na-arụ ọrụ n'onwe ya ruo mgbe arụ ọrụ nyocha niile gwụchara.

Nyocha nke nyocha scIC

Usoro nyocha nke scIC

• Ihe data scIC nwere ike ịdị iche na data biosensor ọkọlọtọ n'ihi mgbagwoju anya okike nke ihe na-eme na sel ejidere na elu mgbawa heliXcyto, nke nwere ike ịkpata mmebi nke sensọgram.
• Ya mere, a na-emesi ike njikwa mma maka onyonyo ewepụtara na usoro nha ọ bụla yana maka sensọ emepụtapụtara na ibe ọdịda nke nyocha akpaghị aka.
• Na mgbakwunye, heliOS na-enye ngwaọrụ iji lebara mmebi nke sensọgram anya n'oge nyocha data ntuziaka scIC.

Usoro nyocha data scIC:

1. Nyocha onyonyo:
   • Nyochaa onyonyo niile ewepụtara n'oge nha (taabụ onyonyo na mpio nnwale).
   • Selụ ole kwụsiri ike na ọnyà ndị ahụ n'oge ntule niile?
   • Kedu ihe bụ ọnọdụ sel? Lelee maka granularity na iguzosi ike n'ezi ihe cell.
   • Ọnyà ndị ahụ hà dị ọcha ma e mechaa nzọụkwụ ọhụrụ?
2. Nleba data efu:
   • Nyochaa data raw maka ntụpọ ntụaka 1 na ntụpọ ntụpọ 2
   • Mgbama nke ọnụ ọgụgụ nkịtị kwesịrị ịdị nso ma ọ bụ karịa karịa mgbama data raw kacha elu.
   • Mgbama a ọ na-alaghachi na ọkwa ntọala na ntụpọ abụọ ahụ, ma ọ bụ enwere ihe akaebe nke njide na-akọwaghị?
3. Nyochaa data ahaziri ahazi:
   • Ịgba ntụtụ a na-awụ elu maka ntụpọ 1 na ntụpọ 2 na-ekpuchi nke ọma?
4. Nyochaa data atụnyere aka:
   • Nzọụkwụ mmụgharị ahụ ọ na-eweghachi mgbama ahụ na ntọala? Ọ bụrụ na ọ bụghị, lelee ma enwere sel ma ọ bụ irighiri ihe dị na ọnyà mgbe emegharịrị site na ịmalitegharịviewna foto.
   • Enwere spikes, ndị na-apụ apụ, ma ọ bụ mmebi iwu ndị ọzọ na akụkụ nke ntụnyere aka? Ọ bụrụ otu a, regharịaview onyonyo iji chọpụta ihe nwere ike ime ma tụlee mgbanwe ntuziaka.
5. Lelee data dabara adaba:
   • Na-eme nyocha ịdị mma anya
   • Ihe dabara adaba ọ na-akọwa n'ụzọ ziri ezi omume nke akụkụ akụkụ ahụ, ma ọ bụ ahịrị dabara adaba na-agafe sensọgram?
   • Ihe dabara adaba ọ na-akọwa n'ụzọ dabara adaba curvature nke data?
   • Ọ bụ amplitude kwekọrọ na data zoro ezo?

Nyocha data akpaghị aka scIC

• Nyocha akpaaka nke scIC na-ahazi data raw na nkata njikwa mma yana data dabara adaba na ọpịpị ole na ole.
  1. Mepee nnwale scIC site na ịpị ugboro abụọ na nnwale ahọpụtara na Ndepụta nnwale
  2. Pịa bọtịnụ nyocha nnukwu anụnụ anụnụ dị na ala nke taabụ nnwale.
  3. Site na usoro nnwale, họrọ nyocha nke ịchọrọ inyocha.
  4. Họrọ Kinetics scIC site na ụdị nyocha dị wee pịa Ọzọ.
     • IHE: Ọ bụrụ na nnwale ahụ ka na-agba ọsọ, naanị nyocha emechara ka a ga-egosi na ndepụta ahụ. Ozugbo ahọpụtara nyocha, windo na-esote ga-egosipụta ụdị nyocha niile dị kpọmkwem na usoro/nyocha eji.
  5. Ọ bụrụ na ọ dị mkpa, hazie nyocha na windo na-esonụ. Ihe nlere nke dabara adaba bụ “Kinetics – Ọkwa ngwụcha efu” yana igbe “Force fit end end level to Zero” na-arụ ọrụ. Wepụ na ihe nlereanya a naanị ma ọ bụrụ na bayoloji ma ọ bụ data chọrọ nkọwa dị mgbagwoju anya karị.
  6. Ozugbo ahazichara nhazi ahụ, pịa Nyochaa ka ịhụ foto ọ bụla ewepụtara, ihe mmetụta sensọ nke raw na emeziri, yana ụkpụrụ kinetics.
• N'ihi mgbagwoju anya nke sampA na-eji ya eme ihe maka nyocha scIC, ihe mmetụta sensọ na-akpata nwere ike inwe mmejọ.
• Iji dozie nke a, a na-emesi njikwa mma nke onyonyo na sensọgram na nyocha akpaghị aka.
• Ọ bụrụ na achọrọ ndozi akwụkwọ ntuziaka ma ọ bụ nyocha miri emi karịa, a na-enye ngwaọrụ maka ndozi ihe n'ime nyocha akwụkwọ ntuziaka Scratchpad.

Nlekọta na mkpochapụ heliXcyto

• Nlekọta heliXcyto dị oké mkpa iji hụ na ngwá ọrụ ahụ dị mma na ogologo ndụ. Nlekọta anya mgbe niile na-agụnye usoro nhicha nke na-egbochi mmetọ ma nọgide na-eguzosi ike n'ezi ihe nke usoro ahụ, na-eme ka nsonaazụ ziri ezi na ịrụ ọrụ dị nro. Nlekọta oge niile dị oke mkpa maka ntụkwasị obi akụrụngwa yana ogo nsonaazụ nnwale.
  mmezi heliXcyto gụnyere usoro isi abụọ: Cyto System Wash, nke enwere ike itinye ya ozugbo n'ime usoro nyocha nyocha, yana Clean & Sleep, nke a na-eme dị ka usoro dị iche iche iji nọgide na-eji ngwa ahụ ozugbo tupu ejiri ya mee ihe n'ogologo abalị gara aga ma ọ bụ mgbe ejighi ya ihe karịrị ụbọchị 2.
• Ha abụọ na-asachapụ sistemu yana dị ọcha na ụra chọrọ ọnụnọ nke heliX® Chip Mmezi na tray mgbawa.

Ịdị ọcha & ihi ụra

• Usoro dị ọcha & ihi ụra bụ maka ihicha na imechi/echekwa ngwaọrụ. Usoro a na-eji mmiri ultrapure na-asachapụ tubing fluidic nke ngwaọrụ heliX® na mbụ wee jiri ethanol 70%. N'ikpeazụ, a na-ekpuchi tubes niile na ikuku.
• Usoro nhicha a bụ akpaaka zuru oke ma were ihe dịka nkeji iri anọ. N'oge usoro ahụ, a na-agbanye ethanol n'ime karama mmiri, ya mere, a ga-atụfu ihe dị na karama ahụ ma emesịa.
• Mgbe Ọcha & Ụra gachara, ngwa ahụ na-abanye n'ọnọdụ ụra na enweghị ike iji ya ruo mgbe emere Teta & Prime. A chọrọ mgbawa mmezi heliX® maka ọsọ abụọ a.
• Mkpa: Iji gbochie mmetọ tube, jide n'aka na ị ga-atụfu ihe dị n'ime karama mmiri (ngwakọta mmiri / ethanol) ma kpochaa ihe mkpofu mgbe usoro nhicha gasịrị.

Emechiri na nchekwa ogologo oge

• Maka ogologo oge nke anaghị eji ya eme ihe, biko wepụ karama ahụ nwere mmiri na ethanol mgbe usoro dị ọcha & ụra si na tray buffer wee hapụ tubing ahụ n'ikuku.
• Wepukwa Chip Mmezi nke heliX® ma chekwaa n'ime akpa mbụ ya. Nke a na-egbochi nlọghachi azụ na mmetọ. Gbanyụọ ngwaọrụ.

Sistemụ ịsa ahụ

• HelXcyto Sistemu Wash nke ukwuu na-asachapụ sistemu microfluidic site na iji Ngwọta Cleaning 3 (CS3) n'ime ihe dịka nkeji iri anọ na ise. A na-eburu CS3 na nkeji abụọtages. Agịga mbụ na sampA na-ejupụta lep ma tinye ya iji wepụ ihe ọ bụla na-emerụ emerụ.
• IHE: Gbaa ịsacha sistemu Cyto opekata mpe kwa ụbọchị mgbe eji ngwaọrụ ahụ. Anyị na-akwado ịgbakwunye usoro ịsa ahụ Cyto na usoro nyocha nke nyocha ma ọ bụrụ na nyocha nke ọ bụla (mgbe ị na-atụgharị na ahịrị cell ọhụrụ ma ọ bụ ọ bụrụ s).ample quality bụ suboptimal) ma ọ bụ na njedebe nke a assay workflow. mgbawa na-edozi mgbanaka mgbe kacha nke 50 Cyto Sistemu washes egosiri dị ka Ntugharị agụta na tray Chip. view nke njikwa ngwaọrụ.

Kpọtụrụ

• Dynamic Biosensors GmbH
• Perchtinger Str. 8/10
• 81379 Munich
• Germany
• Ụgwọ nke ụlọ ọrụ Bruker Scientific LLC
• 40 Manning Road, Manning Park
• Billerica, MA 01821

USA

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• Nkwado ndị teknuzu support.dbs@bruker.com
• www.dynamic-biosensors.com
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• ©2025 Dynamic Biosensors GmbH
• Maka Jiri naanị nyocha. Ọ bụghị maka ojiji na usoro nyocha ụlọ ọgwụ.

FAQ

SIC FAQ

Enwere m ike iji mgbawa heliXcyto ọzọ?

Ee, mgbawa heliXcyto bụ nke enwere ike ijikwa ya ma tụfuo ya. Soro ntuziaka ihicha na nchekwa kwesịrị ekwesị.

Gịnị bụ ebumnobi nke Chip Mmezi?

A na-eji Chip Mmezi ahụ eme ihe maka nhicha, nnwale, na arụrụ ọrụ iji hụ na arụrụ ọrụ nke sistemu kwesịrị ekwesị.

Ugboro ole ka m ga-ehicha ngwaọrụ ahụ?

A na-edobe sistemu mmiri mmiri nke heliXcyto na usoro ịsa ahụ na Cyto System dị ọcha. A na-atụ aro ka ihicha sistemu microfluidic site na Cyto System Wash na Chip Mmezi mgbe nyocha nke ọ bụla (karịsịa mgbe ị na-agbanwe ụdị cell) ma ọ bụ nke kachasị ọhụrụ na njedebe nke usoro nyocha. A ga-etinye usoro dị ọcha&ụra ozugbo tupu ejiri ya mee ogologo oge gara aga (dịka n'ụtụtụ mgbe nnwale abalị gasịrị) ma ọ bụ mgbe heliXcyto agaghị eji ihe karịrị ụbọchị abụọ mechie.

Ogologo oge ole ka a ga-echekwa ihe ngwọta: RB 1 / mmiri / CS 1 / CS 3 / Normalisation ngwọta na ngwaọrụ?

N'ozuzu, tupu nha nke ọ bụla, a ga-enyocha ihe ngwọta niile maka ụda fọdụrụnụ yana maka oke mmiri ozuzo ma ọ bụ mmiri ozuzo. N'ọnọdụ dị otú ahụ, ọ dị mkpa ka a gbanwee ngwọta ahụ ozugbo. A ga-edochi mmiri na RB 1 kwa ụbọchị yana RB 1 ga-enyocha tupu nha ọ bụla. Enwere ike iji ngwọta Normalisation diluted maka ụbọchị 2 n'usoro. Ngwọta nhicha kwụsiri ike maka ihe dịka ụbọchị 3.

Ogologo oge ole ka m ga-eji mgbawa heliXcyto?

Enwere ike ịlele ụbọchị ngwụcha nke mgbawa heliXcyto na taabụ Chip tray na ngalaba ngwaọrụ dị na heliOS. Enweghị nkwa maka iguzosi ike n'ezi ihe mgbawa nwere ike inye mgbe ụbọchị njedebe gachara. Ka o sina dị, ọ bụrụhaala na ọnyà ndị ahụ na-adịgide adịgide, elu ahụ dị ọcha, na nzụlite fluorescence dị n'okpuru nkwụsị nke egosiri na Nnwale Chip, a na-ewere mgbawa dị ka ihe eji eme ihe. Nhicha mgbawa mpụga mgbe niile (A na-atụ aro ka emee ngwa Cleaning Chip CCK-1-1 mgbe ejiri mgbawa mee ogologo oge ndụ mgbawa nke ukwuu.

Ogologo oge ole ka m ga-eji mgbawa mmezi?

Achọrọ ka edochie mgbawa mmezi ahụ mgbe ịsachara sistemu 50 (gụọ dị ka mmeghari na taabụ Chip tray nke njikwa ngwaọrụ na heliOS).

Ugboro ole ka a ga-eme ule mgbawa heliXcyto?

Nnwale Chip na-anakọta ozi gbasara ndabere fluorescence, ịdị ọcha na iguzosi ike n'ezi ihe nke mgbawa tupu ịmalite nnwale ọhụrụ. A ga-eme ya ma ọ dịkarịa ala otu ugboro mgbe a na-eji mgbawa ọhụrụ, ma a na-atụ aro ka eme ya tupu ma ọ bụ na mmalite nke nnwale ọ bụla iji lelee ọkwa nke mgbawa ahụ. Nke a na-enyere aka gbanwee ọnọdụ nha yana adịghị mma na sensọgram ka a chọtara na mgbawa ma ọ bụ ngwaọrụ.

Kedu ihe bụ kemịkalụ akara nyocha?

A na-ejikọta akwa fluorescent na-acha uhie uhie ma ọ bụ akwụkwọ ndụ akwụkwọ ndụ, n'otu n'otu, site na ngwa ntinye aha anyị site na NHS-esters na amines bụ isi na nyocha protein (dịka lysines ma ọ bụ N-terminus). Enwere ike ịchọta nkọwa na ngalaba nchọpụta nsogbu na ngwa heliXcyto Labeling Kit red dye (Labeling Kit red dye 2 CY-LK-R2-1) na heliXcyto Labeling Kit green dye (Labeling Kit green dye CY-LK-G1-1 ) akwụkwọ ntuziaka sitere na anyị. webụlọ ahịa.

Ebee ka ịchọta nzere heliOS na ozi ikike?

A kọwara usoro nke itinye nzere na ozi ikike na ngalaba nrụnye heliOS na ntuziaka heliXcyto sitere na anyị. websaịtị (heliXcyto ndu). Ekwesịrị ịchekwa ozi ikike gị na nchekwa scIC na heliXcyto PC Desktop, nke Ọkachamara Ngwa anyị mepụtara n'oge ọzụzụ.

Gịnị bụ oke mkpali/napụta nke heliXcyto?

Mmasị na-acha uhie uhie bụ 605-625 nm, nmịka (nchọpụta) bụ 655-685 nm. Mmasị na akwụkwọ ndụ akwụkwọ ndụ 490-510 nm, ikuku na-acha akwụkwọ ndụ bụ 525-575 nm.

Ugboro ole ka m ga-atụle otu nnwale ahụ maka ntụkwasị obi ọnụ ọgụgụ?

Iji nweta nkezi ọnụego kinetic nke a pụrụ ịdabere na ya, a na-atụ aro ka emee nkwugharị 3-4 nke nha Kinetics n'ime ọnụ ọgụgụ cell. N'ime usoro data na-agbanwe agbanwe, mkpakọrịta na ọnụ ọgụgụ nkewapụ nke mmeghari kwesịrị ịdị n'ime windo ugboro 2-4.

Kedu ihe mmetụta nke nha scIC?

Oke nchọpụta dị ala n'ihe gbasara okwu ebumnuche dabere na akara nyocha, mana ọ na-abụkarị scIC nwere ike tụọ kinetics dị ole na ole dị ka molekul 1000-10000 kwa cell. Iji mee ka uche dịkwuo elu, ọ dịkarịa ala sel 5 ka a na-atụ aro ka ejide ya, yana nyocha DOL nke 5-10 na ike LED ga-edozi ahụ iji nweta ọnụ ọgụgụ fluorescence kacha.

Kedu oke nchọpụta nke heliXcyto n'ihe gbasara ọnụego kinetic?

Biko rụtụ aka na nkọwa nka nke heliXcyto maka oke teknụzụ kachasị ọhụrụ, nke enwere ike ịhụ na ntuziaka heliXcyto (nduzi heliXcyto). Dissociation mgbe niile Kd: 10 pM ka 1 mM Association ọnụego mgbe niile kon: 1E3 ka 1E7 M-1s-1 Dissociation ọnụego mgbe nile koff: 1E-6 ka 1 s-1 Maka ozi zuru ezu biko lelee anyị heliXcyto ndu si anyị. websaịtị.

Akwụkwọ / akụrụngwa

dynamic BIOSENSORS heliX cyto Sistemụ nyocha nyocha ụlọ nyocha zuru oke [pdf] Akwụkwọ ntuziaka onye ọrụ heliX cyto Sistemụ nyocha ụlọ nyocha zuru oke, heliX cyto, Sistemụ nyocha ụlọ nyocha zuru oke, sistemụ nyocha ụlọ nyocha akpaaka, sistemụ nyocha ụlọ nyocha, sistemu nyocha, sistemụ.

Ntụaka

• Akwụkwọ ntuziaka onye ọrụ