流式常用荧光染料.doc

如何选择流式细胞仪测定常用的荧光染料流式细胞仪测定常用的荧光染料有很多种,我们如何来选择流式细胞仪测定的常用荧光染料呢？ 首先要考虑，他们分子结构不同,激发光谱和发射光谱也不同,选择荧光染料时，必须依据流式细胞仪所配备的激光光源的发射光波长做为重点考虑。发射光波长，如：氩离子气体激光管,它的发射光波488nm； 氦氖离子气体激光管发射光波长633nm；488nm激光光源常用的荧光染料有FITC（异硫氰酸荧光素）； PE（藻红蛋白） PI（碘化丙啶）CY5（化青素） preCP(叶绿素蛋白) ECD(藻红蛋白-得克萨斯红)等。激发光和发射光波长，如下：激发光波长（nm）发射光峰值（nm）FITC488525（绿）PE488575（橙红）PI488630（橙红）ECD488610（红）CY5488675（深红）PreCP488675（深红）流式细胞仪中常用的荧光染料光学特征以及需要的激光器名称Ex波长(nm)Em波长(nm)备注激光器Hoechst 33342 355465DNA染色核酸标记351nm紫外激光器Hoechst 33258365465DAPI372456Indo-1350405细胞内钙离子标记FITC490520标记抗体488nm氩离子激光器R_PE480578PE_CY5480670PE_CY5.5480695PerCP490675Alexa Fluor 488 494517PE_Texas Red 480613Rhodamine 123 500540线粒体标记Fluo-3506526细胞内钙离子标记DiOC6(3)480501内质网标记YOYO-1490510DNA染色核酸标记Propidium Iodide530615Acridine Orange490640APC650660标记抗体633nm氦氖激光器Alexa Fluor 647 650668CY5650667流式细胞仪交流论坛：/forum-181-1.html荧光染料谱染料名称Excitation(nm)Emission(nm)分子量备注信息Reactive and conjugated probesHydroxycoumarin325386331Succinimidyl esterAminocoumarin350445330Succinimidyl esterMethoxycoumarin360410317Succinimidyl esterCascade Blue(375);401423596HydrazidePacific Blue403455406MaleimidePacific Orange403551Lucifer yellow425528NBD466539294NBD-XR-Phycoerythrin (PE)480;565578240 kPE-Cy5 conjugates480;565;650670aka Cychrome, R670, Tri-Color, Quantum RedPE-Cy7 conjugates480;565;743767Red 613480;565613PE-Texas RedPerCP490675Peridinin chlorphyll proteinTruRed490,675695PerCP-Cy5.5 conjugateFluorX494520587GE HealthcareFluorescein495519389FITC; pH sensitiveBODIPY-FL503512TRITC547572444TRITCX-Rhodamine570576548XRITCLissamine Rhodamine B570590Texas Red589615625Sulfonyl chlorideAllophycocyanin (APC)650660104 kAPC-Cy7 conjugates650;755767PharRedAlexa Fluor系列荧光染料Alexa Fluor 350343442410Alexa Fluor 4054014211028Alexa Fluor 430434540702Alexa Fluor 488499519643QY 0.92Alexa Fluor 500503525700Alexa Fluor 514517542714Alexa Fluor 532530555724QY 0.61Alexa Fluor 5465615721079QY 0.79Alexa Fluor 5555535681250QY 0.1Alexa Fluor 568579603792QY 0.69Alexa Fluor 594591618820QY 0.66Alexa Fluor 6106106291285Alexa Fluor 6336326481200Alexa Fluor 6476526681300QY 0.33Alexa Fluor 6606636911100Alexa Fluor 6806807021150Alexa Fluor 7006967191400Alexa Fluor 7507527761300Alexa Fluor 7907828041750Cy系列荧光染料Cy2489506714QY 0.12Cy3(512);550570;(615)767QY 0.15Cy3B558572;(620)658QY 0.67Cy3.5581594;(640)1102QY 0.15Cy5(625);650670792QY 0.28Cy5.56756941128QY 0.23Cy7743767818QY 0.28核酸相关探针染料Hoechst 33342343483616 AT-selective DAPI345455AT-selective Hoechst 33258345478624AT-selective SYTOX Blue431480400DNA Chromomycin A3445575CG-selective Mithramycin445575YOYOthidium Bromide493620394 Acridine Orange503530/640DNA/RNASYTOX Green504523600 DNA TOTO-1, TO-PRO-1509533Vital stain, TOTO: Cyanine Dimer TO-PRO: Cyanine MonomerThiazole Orange510530Propidium Iodide (PI)536617668.4LDS 751543;590712;607472 DNA (543ex/712em), RNA (590ex/607em) 7-AAD5466477-aminoactinomycin D, CG-selectiveSYTOX Orange547570500 DNA TOTO-3, TO-PRO-3642661DRAQ5647681,697413usable excitation down to 488细胞功能探针Indo-1361/330490/4051010AM ester. Low/High Ca+,Fluo-3506526855AM ester. pH 6DCFH50553552927Dichorodihydrofluorescein, oxidized formDHR505534346Dihydrorhodamine 123, oxidized form, light catalyzes oxidationSNARF548/579587/635pH 6/9荧光蛋白QYBRPSY66H360442Y66F360508EBFP3804400.180.27monomerEBFP238344820monomerAzurite38344715monomerGFPuv385508T-Sapphire3995110.602625weak dimerCerulean4334750.622736weak dimermCFP4334750.401364monomerECFP4344770.153CyPet4354770.511859weak dimerY66W436485mKeima-Red4406200.243monomerTagCFP45848029dimerAmCyan14584890.7529tetramermTFP146249254dimerS65A471504Midoriishi Cyan4724950.925dimerWild Type GFP396,47550826k0.77S65C479507TurboGFP48250226 k0.5337dimerTagGFP48250534monomerS65L484510Emerald4875090.68390.69weak dimer; (Invitrogen)S65T488511EGFP48850726k 0.6034174weak dimerAzami Green4925050.7441monomerZsGreen1493505105k0.9140tetramerTagYFP50852447monomerEYFP51452726k0.615160weak dimerTopaz51452757monomerVenus5155280.575315weak dimermCitrine5165290.765949monomerYPet5175300.778049weak dimerTurboYFP52553826 k0.531.65dimerZsYellow15295390.6513tetramerKusabira Orange5485590.6031monomermOrange5485620.69499monomermKO5485590.6031122monomerTurboRFP55357426 k0.6762dimertdTomato5545810.699598tandem dimerTagRFP55558450monomerDsRed monomer55658628k0.13.516monomerDsRed2 (RFP)563582110k0.5524mStrawberry5745960.292615monomerTurboFP60257460226 k0.3526dimerAsRed2576592110k0.2113tetramermRFP158460730k0.25monomerJ-Red5846100.208.813dimermCherry5876100.221696monomerHcRed158861852k0.03