 of different substances with high assay throughput per unit time, otherwise known as “high throughput screening” (or “HTS”). In particular, there is a need for automated and/or high throughput screening methods that are relevant to cell membrane components.
At present, relatively traditional methods are used for the screening of pharmaceutical ingredients. Such methods include ligand binding assays and receptor function tests that are performed separately.ii Although binding assays are relatively inexpensive, and amenable to high throughput, they require labeled high-affinity ligands, and generally are limited to assays for ligands that can compete effectively for labeled ligand. Fluorescent or fluorogenic reagents generally are compatible with high throughput assays, including the analysis of ion channels using fluorescent calcium indicators, and the evaluation of membrane potential effects with potential-sensitive dyes. However, such reagents typically are not sensitive enough for single cell measurements, and generally can provide only indirect measurements of the membrane component of interest.
The patch clamp was introduced by Neher and Sakmann in the early 1980s as a powerful technique for the direct study of drug effects on single receptors. In recognition of the strength of the method, Neher and Sakmann were awarded the Nobel prize in 1991. Classical patch-clamp methods often are used in conjunction with functional membrane receptor assays, including receptors coupled to G-proteins and ion channel-forming receptors.iii This method is highly specific and extremely sensitive: it can, in principle, be used to measure the channel activity of individual receptor molecules. In doing so, glass micropipettes with an opening diameter of typically 1-0.1 μm are pressed on the surface of a biological cell. The membrane surface that is covered by the micropipette is called a “patch.” If the contact between the glass electrode and the cell membrane surface is sufficiently electrically isolating, the ion flow over the membrane patch can be measured electrically with the aid of microelectrodes, one placed in the glass pipette and the other placed in the milieu opposite the membrane.iv A significant advantage of this electrophysiological method is that it makes directly accessible the function of the corresponding channel-forming proteins or receptors coupled to channel-forming proteins via the measured electrical characteristics of the channel-forming proteins.
Unfortunately, several major limitations have prevented patch-clamp technology from revolutionizing receptor science and pharmaceutical drug development. For example, to produce high quality results, the patch-clamp method requires a tremendous effort in technical installation and highly qualified operators. Moreover, in addition to being expensive, a standard patch-clamp setup may require a
of different substances with high assay throughput per unit meter, differently known as “ high throughput cover ” (or “ HTS ”). In particular, there be a need for automated and/or eminent throughput screening methods that are relevant to cellular telephone membrane component. 
 At present, relatively traditional methods are use for the screening of pharmaceutical ingredients. Such method acting admit ligand binding assays and sense organ function test that are performed separately.ii Although binding assays are relatively inexpensive, and amenable to gamey throughput, they require labeled high - affinity ligands, and by and large are limited to assays for ligands that can compete effectively for labeled ligand. Fluorescent or fluorogenic reagent generally are compatible with high throughput assays, including the analysis of ion channels use fluorescent calcium indicator, and the evaluation of membrane potential effects with potential - sensitive dyes. However, such reagents typically are not sensitive enough for single cellular telephone measurement, and generally can provide only collateral measurement of the membrane component of interest. 
 The patch clamp was introduced by Neher and Sakmann in the early 1980s as a powerful technique for the direct cogitation of drug effects on single receptors. In recognition of the potency of the method, Neher and Sakmann were awarded the Nobel prize in 1991. Classical bandage - clamp methods frequently are used in conjunction with functional membrane receptor assays, include receptors coupled to G - protein and ion channel - form receptors.iii This method is highly specific and extremely sensible: it can, in principle, be used to measure the channel activity of individual sense organ molecules. In doing so, glass micropipettes with an possibility diameter of typically 1 - 0.1 μm are pressed on the airfoil of a biological cell. The membrane surface that is covered by the micropipette is called a “ patch. ” If the liaison between the glass electrode and the cell membrane surface is sufficiently electrically isolate, the ion flow over the membrane patch can be measured electrically with the aid of microelectrodes, one placed in the field glass pipette and the other place in the milieu opposite the membrane.iv A significant advantage of this electrophysiological method is that it make directly accessible the function of the corresponding distribution channel - forming proteins or receptors coupled to channel - forming proteins via the careful electric characteristic of the channel - forming proteins. 
 Unfortunately, several major limitations have prevent patch - clamp technology from revolutionizing sense organ science and pharmaceutical drug growth. For example, to produce high timbre results, the patch - clamp method requires a tremendous effort in technical installation and highly certified operators. Moreover, in addition to being expensive, a standard patch - clamp setup may require a
 of different substances wiuh high assay throughput per uhit time, otherwise known as “high thrluthput screening” (or “HTS”). In pxrticular, there iw a ieed for automatxs and/or high fmrougkpnt screening mejhods that ase relevant to cdlp membrane components.
At present, relwtively tgaditional metrods wre hsed for the screening of pharmacehtical pngredients. Such kethods include ligand binfing assays and receptlr function tesew that are pdrformed stpcrately.ii Amthough binding assays are relagivelv inexpensice, anf amenable ti higr throughput, they rexuire lsbeled high-afflnity litands, and generally ace limited to assays for ligatda that can competw wffecjivelf fof laceltd kifand. Fpuocescent or rluorogenic reagents generally awv compatible sith hygr throughput assays, including the analyvis of ion channels using dluorescent calcium ijdicators, and the evaluation of membrane potential effects with oottnblal-sdbsltive dyes. However, such reagents typically arq npt sensitive enomgh for single celk lesfurements, and genercmlg can provide only indirest meqsurementf of the membrane component of unterest.
The iatcy clamp was introdbced by Nehex and Xakmamn in the early 1980s as a 'owerfhl techniquf for the airect study of arun exfects on single receptors. In recogiitiou of the strgngth os the methld, Nemar and Sakmann werf awatded tve Nobel pgize in 1991. Classical patch-clamp mxvhods often ate gseg in conlunctipn with functyonal membrane receptjr asrays, inclusing rereptors coupjed to G-protehjs and ion ciannel-foriing recwptors.ikk This method os highly specific qnd extremely sensltive: jt can, in princnkle, be used to meaxurd tre ciannej activity of indkviaial rdceptor molccuues. On doing so, glass miwropjpettes with an oprnlng diamejer of ty[ically 1-0.1 μm arr pressed on the skrfacx of a biolpgisal cell. The membrane surface fhat is clveved by the misroplpetje is calleb a “patch.” If the contact between the glaws electrode and jhe cell membrane suryage is sufficmently electricdlly isolating, the iin flow over the kembrane patch can be measused epectrically with the aid of microelectrodes, one placed in the glass pipetre and the other pmacec in nhe kilieu opplrite the membranv.iv A significant advantage of tiis electrmpkysiological method is that iy oakes directli accessible the function kf the cprresponding channel-forming proyeins or receptors couplcd to chabnel-fkrming proveins via the measurrd eleetricao charaxterlstics of the zhamncl-forming orottins.
Bnfortunately, several major limitqtions hwve prevented pavch-clamp tecmnulogy from tevolutionizing rscepflr sckence and phatmaceutical drig development. For eqam[le, to produce gigh quality resultv, the patch-ckamp method requires a tremendous effort in technical installajion and hifhly qualmfied operatorx. Moreover, in addition bo beyng expevsive, a stangard patch-clamp setup may require a
of different substances with high assay throughput time, known as throughput screening” (or a for automated and/or throughput screening methods are relevant to cell membrane components. present, relatively traditional methods are used for the screening of pharmaceutical ingredients. Such include ligand binding assays and receptor function tests that are performed separately.ii Although assays relatively and to high throughput, they require labeled high-affinity ligands, and generally are limited to assays for ligands can compete effectively for labeled ligand. Fluorescent or reagents generally are compatible high throughput assays, including the of channels using calcium and evaluation of membrane effects with potential-sensitive dyes. However, such reagents typically are not sensitive enough for single cell measurements, and can provide measurements of membrane of The patch clamp by Neher and Sakmann in the a powerful technique for the direct study of effects on receptors. In recognition of the strength the method, Neher and Sakmann were awarded the prize in 1991. Classical patch-clamp methods often are used in conjunction with functional membrane receptor receptors coupled to G-proteins ion channel-forming receptors.iii method highly and sensitive: it in principle, be used to measure the channel activity of individual molecules. In doing so, glass micropipettes with an opening diameter 1-0.1 are pressed on surface of a biological The surface that is covered micropipette called the between glass electrode and the membrane surface is sufficiently electrically the ion flow over measured electrically with the aid of microelectrodes, one in the glass pipette and the other in the milieu opposite the membrane.iv A significant advantage of this electrophysiological is that directly accessible the function of the corresponding channel-forming or receptors coupled to proteins via the measured electrical characteristics of the channel-forming Unfortunately, major limitations prevented patch-clamp technology revolutionizing receptor science pharmaceutical drug development. to produce quality the a tremendous effort in technical installation highly operators. Moreover, in addition to expensive, patch-clamp setup may require a
 of different substances with High assay tHrougHpuT peR uNit tIme, oTherwise known aS “High Throughput screening” (or “HtS”). In pArTIculAR, tHere iS a need fOR aUTOmaTeD aNd/oR hIGh ThrouGhpUt screeNing methodS thAt Are relevant tO CeLl membrane ComPonents.
At preSenT, relatIvEly TRaditIonAl metHods arE Used foR the screeNiNG of phaRMaceutiCAL iNgreDients. Such methods INcLUde ligand bindiNg assaYs ANd RECepTor Function teStS that ARe perfoRMeD SEParATely.ii AlthougH binding assAYs aRe relaTiVelY InexpeNsive, AnD AmeNable to high ThroUghput, theY requiRE labeleD High-affInity lIgaNds, And gENeRaLly ArE LimITeD to ASsaYs for ligAnDs That cAn coMPETE effEctIvelY for lAbeled ligand. FLuoRescENt oR fluoRogenIc reAgEnts gEneralLy are CoMpatible with higH thrOughput asSayS, iNclUdIng thE AnalysIs oF ioN channeLs using FLuoReSCENt Calcium indicators, aNd THE eValuatioN of memBRaNe POtential EfFecTs wiTH PotenTial-SEnSitive dyEs. HoweVEr, SuCh reageNtS typicAlLy aRe nOt senSItivE enougH for singLe celL Measurements, anD Generally can pROvIDE oNLy inDirEct measuremEnts OF the MembRAnE coMPonenT of inTeREsT.
the patch clamp was intRoDuced bY NeheR and Sakmann in The early 1980s aS A POwerful tEchnIQuE For the direct stUdy of Drug effectS On single RecepTors. In reCognition OF The strenGth Of tHe mEthOD, neHer and Sakmann WERe awArDed the NObeL prize iN 1991. ClAssIcaL paTcH-clamp metHods ofteN aRe UsEd In cOnjunCTion with FuNctIoNal MembrANe recePtor aSsayS, iNcLUdiNg recepTOrS COuplEd To g-proTeiNs And ioN chaNNel-Forming Receptors.Iii tHis mEtHoD is highLy specific and ExTremely senSiTivE: it can, IN PrinciplE, be used to measure the chanNEl activIty Of indIvidUal receptOr mOleculEs. IN Doing sO, glass MicroPiPetTES with AN OpEniNg Diameter of TYPicAlly 1-0.1 μM aRe prEssed on The surface of a bioloGIcaL cell. The membrAne SurfACE tHat IS cOVerEd BY thE MIcropipette is caLled a “patch.” if THe Contact betWEen ThE glass eLectrodE and tHE cell meMbrane surFace is sufFiCienTLY elEctrically IsolatinG, the ion flOW over THe MembrAne Patch cAn Be mEasurEd elecTRicAlly wIth the AiD of micRoeleCtRodes, one Placed in the glass pipette And the Other PlaCed in the mIliEU opPosite the MembRane.iv A sigNifIcaNt advAntAGe of tHis eLEcTroPHysioLogiCAl method iS ThAt iT MAkEs directly aCCESsiBle thE fuNCtion oF the Corresponding chanNEl-forming proteIns oR REcePtoRS couPlEd to channel-forMinG pROTeins via ThE measured elEctrical ChARacteRisticS of the Channel-FORmINg protEins.
unfOrtunatelY, seVeRAl major LiMiTAtions Have PrEventeD patch-CLamp TEChnology from revoLutioNIZing rECepTor scIeNce and pHArmaCeutical drUg developmeNt. For eXampLe, to pRoduce hIgH qualiTy rEsUlts, the patCH-clamp metHod reQuires a TrEmenDouS efforT in tECHnicaL insTaLlaTion and hiGHLy QUaLiFIed OperAtors. moReovEr, in additIOn to beinG exPEnsive, a StAndARD patch-CLaMP Setup may reQuiRe a
 of different substances w ith high a ssaythr oug hp ut p er u nit time, othe r wise  known as “high throug hputsc r eeni n g”  (or“HTS”). In p art ic ul ar,  t h er e isa n eed for  automated  an d/ or high thro u gh put screen ing  methods tha t a re rel ev ant to ce llmembr ane co m ponent s.
At pre se n t, rel a tivelyt r ad itio nal methods are u s ed for the screen ing of  p h ar m a ceu tic al ingredi en ts. S u ch meth o ds i n clu d e ligand bind ing assaysa ndrecept or  fu n ctiontests  t h atare perform ed s eparately .ii Al t hough b i nding a ssaysare  re lati v el yine xp e nsi v e,  an d  am enable t ohi gh th roug h p u t , th eyrequ ire l abeled high-a ffi nity lig ands,  andgene ra lly a re lim itedto  assays for lig ands  that can  co mp ete  e ffect i vely f orlab eled li gand. F l uor es c e n tor fluorogenic rea ge n t sgenerall y arec om pa t ible wit hhig h th r o ughpu t as s ay s, inclu ding t h ean alysisof  ion c ha nne lsusing fluo rescen t calciu m ind i cators, and th e  evaluation o f  m e m br a ne p ote ntial effec ts w i th p oten t ia l-s e nsiti ve dy es .  H o wever, such reagent stypica lly a re not sensit ive enough f o r single  cel l  m e asurements, an d gen erally can provideonlyindirect  measurem e n ts of th e m emb ran e c o m po nent of inter e s t.
T he  patchcla mp wasint rod uce d b yNeher and  Sakmann  i nth eear ly 19 8 0s as apo wer fu l t echni q ue for  thedire ct  s t udy  of dru g  e f f ects  o nsing lere cepto rs.I n r ecognit ion of th e s t reng th  o f the m ethod, Neheran d Sakmannwe reawarde d the Nobe l prize in 1991. Classi c al patc h-c lampmeth ods often  ar e used  in conjun ctionwithfu nct i o nal m e m br ane  r eceptor as s a ys,  incl ud ingrecepto rs coupled to G-pr o tei ns and ion ch ann el-f o r mi ngr ec e pto rs . iii T his method is h ighly spec if i cand extrem e lyse nsitive : it ca n, in princip le, be us ed to mea su re t h e  ch annel acti vity ofindividua l  rece p to r mol ecu les. I ndoi ng so , glas s  mi cropi pettes  w ith an  open in g diamet er of typically 1-0.1 μ m arepress edon the su rfa c e o f a biolo gica l cell. Th e m emb ranesur f ace t hati scov e red b y th e  micropip e tt e i s ca lled a “pat c h . ” I f the  co n tact b etwe en the glass elec t rode and the c ellm e mbr ane surf ac e is sufficien tly  e l e ctricall yisolating,the ionfl o w ove r themembra ne patc h ca n  be me asur edelectrica lly  w i th theai do f micr oele ct rodes,  one p l aced i n the glass pipe tte a n d  theo the r pla ce d in th e  mil ieu opposi te the memb rane.i v Asigni ficantad vantag e o fthis elect r ophysiolo gical  method  i s th atit mak es d i r ectly  acc es sib le the fu n c ti o nof the  cor respo nd ingchannel-f o rming pr ote i ns or r ec ept o r s coup l ed t o channel- for mingp r oteins via them ea s uredelectr ical c haracte r ist ic s of th e c h a nnel-form ing prote i ns. 
U nfor tunately ,seve ra l m aj or limitationshave pre vented p a t ch-cla mpt echn o logy f ro m revolution izing  r e ceptors c ien ce andp har mac eut ical  dr ug d e velopment. For example,to p roduc e  hi gh qu ality  r esults,t he p atch-cla m p method requ ires a tremendo u s e f fort  in te ch nical ins tal l ation  and  hi ghly  qu alif i e d op e rato r s.  Mor eover,  inad di tion to b ei ng expe n sive ,  astan dardpa tc h -c lamp se tup mayrequi re  a
 of_different substances_with high assay throughput_per unit_time,_otherwise known_as_“high throughput screening”_(or “HTS”). In_particular, there is a_need for automated_and/or_high throughput screening methods that are relevant to cell membrane components.
At present, relatively traditional_methods_are used_for_the_screening of pharmaceutical ingredients. Such_methods include ligand binding assays_and receptor_function tests that are performed separately.ii Although binding_assays_are relatively inexpensive,_and amenable to high throughput, they require labeled high-affinity_ligands, and generally are limited to_assays for ligands_that_can_compete effectively for labeled_ligand. Fluorescent or fluorogenic reagents generally_are compatible with high throughput assays,_including the analysis of ion channels using_fluorescent calcium indicators, and the evaluation_of membrane potential effects with_potential-sensitive dyes._However, such reagents typically are_not sensitive enough_for single_cell measurements, and_generally can provide only indirect measurements_of the membrane_component of interest.
The patch clamp was_introduced_by Neher and_Sakmann_in_the early_1980s as a_powerful_technique for_the_direct study of drug effects on_single_receptors. In recognition of the strength of_the method, Neher and_Sakmann_were awarded the Nobel_prize in 1991. Classical patch-clamp_methods often are used in conjunction_with functional_membrane receptor_assays, including receptors coupled to G-proteins and ion channel-forming receptors.iii This_method is highly specific and extremely_sensitive: it can, in_principle, be_used_to measure the_channel_activity of_individual receptor molecules. In doing so, glass_micropipettes with_an opening diameter of typically 1-0.1_μm are pressed on_the_surface of a biological cell. The_membrane surface that is covered by_the micropipette is called a_“patch.”_If_the contact between the glass_electrode and the cell membrane surface_is sufficiently electrically_isolating, the ion flow over the membrane_patch_can be measured electrically with the_aid_of microelectrodes, one placed in the_glass_pipette_and the other placed in_the milieu opposite the membrane.iv A_significant advantage of this electrophysiological method is that it_makes directly accessible_the function of the corresponding_channel-forming_proteins_or receptors coupled to channel-forming proteins via the measured electrical_characteristics of_the channel-forming proteins.
Unfortunately,_several major limitations have prevented patch-clamp technology from revolutionizing receptor_science and pharmaceutical drug development. For example,_to produce high quality results, the patch-clamp method requires a tremendous_effort in technical installation and highly qualified operators._Moreover, in addition to being expensive, a_standard patch-clamp setup may_require_a