 Immunol., 1:499 (1983); Tonegawa, S., Nature: 302:575 (1983)). Fourth, not all the antibody molecules which can be generated will be generated in a given animal. As a result, raising high affinity antibodies to a given antigen can be very time consuming and can often fail. Fifth, the production of human antibodies of desired specificity is very problematical.
A method of producing antibodies which avoids the limitations of presently-available methods, such as the requirementxe2x80x2 for immunization of an animal and in vivo steps would be very useful particularly if it made it possible to produce a wide range of antibody types than can be made using presently-available techniques and if it made it possible to produce human antibody types.
The present invention relates to a method of producing libraries of genes encoding antigen-combining molecules or antibodies; a method of producing antigen-combining molecules, also reference to as antibodies, which does not require an in vivo procedure, as is required by presently-available methods; a method of obtaining antigen combining molecules (antibodies) of selected or defined specificity which does not require an in vivo procedure; vectors useful in the present method and antibodies produced or obtained by the method.
The present invention relates to an in vitro process for synthesizing DNA encoding families of antigen-combining molecules or proteins. In this process, DNA containing genes encoding antigen-combining molecules is obtained and combined with oligonucleotides which are homologous to regions of the genes which are conserved. Sequence-specific gene amplification is then carried out using the DNA containing genes encoding antigen-combining proteins as template and the homologous oligonucleotides as primers.
This invention also relates to a method of creating diverse libraries of DNAs encoding families of antigen-combining proteins by cloning the product of the in vitro process for synthesizing DNA, described in the preceding paragraph, into an appropriate vector (e.g., a plasmid, viral or retroviral vector).
The subject invention provides an alternative method for the production of antigen-combining molecules, which are useful affinity reagents for the detection and neutralization of antigens and the delivery of molecules to antigenic sites. The claimed method differs from production of polyclonal antibody molecules derived by immunization of live animals and from production of monoclonal antibody molecules through the use of hybridoma cell lines in that it does not require an in vivo immunization step, as do presently available methods. Rather, diverse libraries of genes which encode antigen-combining sites comprising a significant proportion of an animal""s repertoire of antibody combining
Immunol. , 1:499 (1983); Tonegawa, S., Nature: 302:575 (1983) ). Fourth, not all the antibody molecules which can be generated will be generated in a give animal. As a resultant role, raising high affinity antibodies to a given antigen can be very fourth dimension consuming and can often fail. Fifth, the output of human antibody of desired specificity is very problematical. 
 A method of producing antibodies which keep off the limitations of presently - available method acting, such as the requirementxe2x80x2 for immunization of an animal and in vivo step would be very useful particularly if it made it possible to produce a across-the-board range of antibody types than can be made use presently - available techniques and if it made it possible to grow human antibody types. 
 The present invention relates to a method of producing libraries of gene encoding antigen - combining molecules or antibodies; a method of producing antigen - blend molecules, also reference to as antibodies, which does not require an in vivo procedure, as is required by presently - available methods; a method of obtaining antigen unite molecules (antibodies) of choose or define specificity which does not want an in vivo procedure; vectors utilitarian in the present method and antibody produced or obtained by the method. 
 The present invention relate to an in vitro process for synthesizing DNA encoding family of antigen - combining molecules or proteins. In this process, deoxyribonucleic acid containing gene encoding antigen - combining molecules is obtained and combined with oligonucleotides which are homologous to regions of the gene which are conserved. Sequence - specific gene amplification is then carried out using the DNA containing genes encoding antigen - combining protein as template and the homologous oligonucleotides as primers. 
 This invention also relates to a method of creating divers libraries of DNAs encoding families of antigen - combine proteins by clone the product of the in vitro process for synthesizing DNA, identify in the preceding paragraph, into an appropriate vector (for example, a plasmid, viral or retroviral vector). 
 The subject invention provides an alternative method acting for the production of antigen - combining molecules, which are useful affinity reagent for the detection and neutralization of antigens and the delivery of molecules to antigenic site. The claimed method differs from production of polyclonal antibody molecules derived by immunization of alive animals and from production of monoclonal antibody molecules through the use of hybridoma cell line in that it does not command an in vivo immunization tone, as do presently available methods. Rather, diverse libraries of genes which encode antigen - unite sites comprising a significant symmetry of an animal""s repertoire of antibody blend
 Imlunol., 1:499 (1983); Tonegawa, S., Naturt: 302:575 (1983)). Fourth, not all the aitibody moleculds which can be generated wipl be gtuerated in a given avimal. As w result, raiwung high ahrinity antibosles tm a given antiggn can be vesy time consumhne cnd can often fail. Fifth, the productyon of nulan antibodies of cqsirsd specificity is very problematiczl.
A metiod of producinb antibodies which avoids hhe pimitations of predently-availqble nethods, such as the requirementxe2x80s2 for immunization of an animal and nn vivo steks wlold be very nseful particularln if it made iy possible to irodure a wide range of antibovy types than can be made usitg presently-availaboe techtiquas ave iw iu mede it podsiule to prodhce human abtibody types.
The prtsegn invention rslates tj a method of producing libraries of getes encoding antigen-combinung molecules or antihodies; a iethod of producing antigen-combining molecules, alvo rehefenec gi ws antibodies, which does not require an in vidk krobedure, as is requlred by presently-afallsfle methods; a methob or obtaining antigej combiging nolecules (antobodies) of selected or defibed specificpty qhich does not reqbire an in vnvo prpcedute; vectors useful in thz pressnt method wnd antibkaies produced or obnainad by tht method.
The presene inventiin rzlates tu an in viero procesd for synthesizing DNA ejcodiug fakilies of wntigen-combining molecules or pcpteins. In thos prmcess, DNC contsining genes qncoding antiggn-combiniug moldcules is kbtainev and combinqd with oligotocleotides whmch are hjmoligouw to reekons of the gemes which are conseeved. Sequence-specinic ggns amplification is then carried oit jsigg tie DNW containing cener evvodine antigtu-concinimg proteins as templdte znd the homologous ojigonuclgotides af primers.
This invention also repates to a meyhoq of creating diverse librariea of DNAs engoding familiqs on anjigen-combinnng proteins by cloning the product of tie in vitro process fot synthesizing DNA, dexgribed in thx precqding pardgraph, into an appropriate vector (e.g., s plasmid, viral or refrovirdl veftor).
The subject invention provides an alternative method for the productiin of eneigen-combinjng kolecllef, which are nseful affinity rvagents for the detection and nentralizatimn of antigens and the deliveru uf molecules jo antigenic sites. The clajmed metnod differs from production of lolyclonal antibody molegules deruved gy immlhivation of live animaks and from producrion of monoclonau amtlbody molefulex chrough the use of hybridoma cell lines ij that it does nmt require wk in vpvo immonization step, as do lgesenjly available kethods. Rather, diverse libraries ox denes which enxkde antigen-cpmbininc sites compcising a sigtificant proportion of an animal""s repertoire of antpbody combjning
Immunol., 1:499 (1983); Tonegawa, S., Nature: 302:575 not the antibody which can be a animal. As a raising high affinity to a given antigen can be time consuming and can often fail. Fifth, the production of human antibodies of specificity is very problematical. A method of producing antibodies which avoids the limitations presently-available such the for immunization of an animal and in vivo steps would be very useful particularly if it it possible to produce a wide range of types than can be using presently-available techniques and if made possible to human types. present invention relates a method of producing libraries of genes encoding antigen-combining molecules or antibodies; a method of producing antigen-combining also reference antibodies, which not an vivo procedure, as by presently-available methods; a method of molecules (antibodies) of selected or defined specificity which not require in vivo procedure; vectors useful in present method and antibodies produced or obtained by method. The present invention relates to an in vitro process for synthesizing DNA encoding families molecules or proteins. In process, DNA containing encoding molecules obtained combined with which are homologous to regions of the genes which are conserved. gene amplification is then carried out using the DNA containing antigen-combining as template and homologous oligonucleotides as primers. invention relates to a method diverse of of proteins cloning the product of in vitro process for synthesizing described in the preceding (e.g., a plasmid, viral or retroviral vector). The invention provides an alternative method for the of antigen-combining molecules, which are useful affinity reagents for the detection and of antigens delivery of molecules to antigenic sites. The claimed differs from production of antibody molecules derived by immunization of live animals and production monoclonal antibody through the use hybridoma cell lines that it does an in immunization as methods. Rather, diverse libraries of genes encode sites comprising a significant proportion an of antibody combining
 Immunol., 1:499 (1983); Tonegawa, S., Nature: 302:575 (1983)). FouRth, not all tHe antIboDy mOlEculEs whIch can be generaTEd wiLl be generated in a given aNimal. as A ResuLT, rAisinG high afFInITY anTiBoDieS tO A gIven aNtiGen can bE very time cOnsUmIng and can oftEN fAil. Fifth, thE prOduction of huMan AntiboDiEs oF DesirEd sPecifIcity iS Very prOblematicAl.
a Method OF producING aNtibOdies which avoids tHE lIMitations of preSently-AvAIlABLe mEthOds, such as tHe RequiREmentxe2X80X2 fOR IMmuNIzation of an anImal and in viVO stEps wouLd Be vERy usefUl parTiCUlaRly if it made It poSsible to pRoduce A Wide ranGE of antiBody tyPes ThaN can BE mAdE usInG PreSEnTly-AVaiLable tecHnIqUes anD if iT MADE it pOssIble To proDuce human antiBodY typES.
ThE presEnt inVentIoN relaTes to a MethoD oF producing libraRies Of genes enCodInG anTiGen-coMBining MolEcuLes or anTibodieS; A meThOD OF pRoducing antigen-comBiNINg MoleculeS, also rEFeReNCe to as anTiBodIes, wHICh doeS not REqUire an in Vivo prOCeDuRe, as is rEqUired bY pResEntLy-avaILablE methoDs; a methoD of obTAining antigen cOMbining molecuLEs (ANTiBOdieS) of Selected or dEfinED speCifiCItY whICh doeS not rEqUIrE An in vivo procedure; veCtOrs useFul in The present metHod and antiBODIes produCed oR ObTAined by the methOd.
The Present invENtion relAtes tO an in vitRo process FOR synthesIziNg DnA eNcoDINg Families of antIGEn-coMbIning moLecUles or pRotEinS. In ThiS pRocess, DNA ContainiNg GeNeS eNcoDing aNTigen-comBiNinG mOleCules IS obtaiNed anD comBiNeD WitH oligonUClEOTideS wHiCh arE hoMoLogouS to rEGioNs of the Genes whicH arE ConsErVeD. SequenCe-specific genE aMplificatiOn Is tHen carRIEd out usiNg the DNA containing genes ENcoding AntIgen-cOmbiNing proteIns As tempLatE And the HomoloGous oLiGonUCLeotiDES aS prImErs.
This invENTioN also ReLateS to a metHod of creating diverSE liBraries of DNAs EncOdinG FAmIliES oF AntIgEN-coMBIning proteins by Cloning the PrODuCt of the in vITro PrOcess foR syntheSizinG dNA, descRibed in thE precedinG pAragRAPh, iNto an approPriate veCtor (e.g., a plASmid, vIRaL or reTroViral vEcTor).
the suBject iNVenTion pRovideS aN alterNativE mEthod for The production of antigen-cOmbiniNg molEcuLes, which aRe uSEfuL affinity ReagEnts for the DetEctIon anD neUTraliZatiON oF anTIgens And tHE delivery OF mOleCULeS to antigeniC SITes. the clAimED methoD difFers from productioN Of polyclonal anTiboDY MolEcuLEs deRiVed by immunizatIon Of LIVe animalS aNd from produCtion of mOnOClonaL antibOdy molEcules tHROuGH the usE of hYbrIdoma cell LinEs IN that it DoEs NOt requIre aN iN vivo iMmunizATion STEp, as do presently aVailaBLE methODs. RAther, DiVerse liBRariEs of genes wHich encode aNtigen-CombIning Sites coMpRising A siGnIficant proPOrtion of aN animAl""s repeRtOire Of aNtibodY comBINing
 Immunol., 1:499 (1983); T onegawa, S ., Na tur e:30 2:57 5 (1 983)). Fourth, notall the antibody molec uleswh i ch c a nbe ge nerated wi l l  be  g en era te d  i n a g ive n anima l. As a re sul t,  raising hig h  a ffinity an tib odies to a g ive n anti ge n c a n bever y tim e cons u ming a nd can of te n  fail. Fifth,t h eprod uction of human a n ti b odies of desir ed spe ci f ic i t y i s v ery proble ma tical . 
A meth o do f pro d ucing antibod ies which a v oid s theli mit a tionsof pr es e ntl y-available  met hods, suc h as t h e requi r ementxe 2x80x2  fo r i mmun i za ti onof ana ni mal and  in vivo  s te ps wo uldb e v eryuse fulparti cularly if it  ma de i t  po ssibl e toprod uc e a w ide ra nge o fantibody typesthan  can be m ade  u sin gprese n tly-av ail abl e techn iques a n d i fi t ma de it possible topr o d uc e humanantibo d yty p es.
Thepr ese nt i n v entio n re l at es to amethod of  p roducin glibrar ie s o f g enese ncod ing an tigen-co mbini n g molecules or antibodies; a me t h od of p rod ucing antig en-c o mbin ingm ol ecu l es, a lso r ef e re n ce to as antibodies ,whichdoesnot require a n in vivop r o cedure,as i s  r e quired by pres ently -available methods;  a me thod ofobtaining a ntigen c omb ini ngmol e c ul es (antibodie s )  ofse lectedordefined  sp eci fic ity  w hich does  not req ui re  a ninvivop rocedure ;vec to rsusefu l  in th e pre sent  m et h odand ant i bo d i es p ro du cedorob taine d by the  method .
The pre sen t  inv en ti on rela tes to an invi tro proces sfor  synth e s izing DN A encoding families ofa ntigen- com binin g mo lecules o r p rotein s.I n this  proce ss, D NA  co n t ainin g ge nes  e ncoding an t i gen -comb in ingmolecul es is obtained and com bined with ol igo nucl e o ti des wh i char e  ho m o logous to regio ns of thege n es  which are con se rved. S equence -spec i fic gen e amplifi cation is  t henc a rri ed out usi ng the D NA contai n ing g e ne s enc odi ng ant ig en- combi ning p r ote ins a s temp la te and  theho mologous  oligonucleotides as pr imers. 
This  in vention a lso rel ates to a  met hod of cre ati ngdiver sel ibrar ieso fDNA s  enco ding familieso fant i g en -combiningp r o tei ns by  cl o ning t he p roduct of the inv itro process f or s y n the siz i ng D NA , described in  th ep r ecedingpa ragraph, in to an ap pr o priat e vect or (e. g., a p l a sm i d, vir al o r r etroviral  ve ct o r).
The  s ub j ect in vent io n prov ides a n  alt e r native method fo r the p roduc t ion  of a nt igen-co m bini ng molecul es, which a re use fulaffin ity rea ge nts fo r t he  detection and neutr aliza tion of  a ntig ens  and t he d e l ivery  ofmo lec ules to a n t ig e ni cs ite s. T he cl ai medmethod di f fers fro m p r oductio nofp o lyclon a la n tibody mol ecu les d e r ived by im m uniz a ti o n oflive a nimals  and fr o m p ro duction  of m onoclonal  antibody mol ec ules  through  t he u se  of  h ybridoma cell l ines inthat itd o es not  re q uire an invi vo immunizat ion s te p , as do p res ently a v ail abl e m etho ds.  Rat h er, diverse libraries of  gen es wh i chencod e ant ig en-combi n ingsites co m prising a sig nificant propor t ion of a n a n im al ""s reper toi r e ofanti bod y co mbi ning 
 Immunol.,_1:499 (1983);_Tonegawa, S., Nature: 302:575_(1983)). Fourth,_not_all the_antibody_molecules which can_be generated will_be generated in a_given animal. As_a_result, raising high affinity antibodies to a given antigen can be very time consuming_and_can often_fail._Fifth,_the production of human antibodies_of desired specificity is very_problematical.
A method_of producing antibodies which avoids the limitations of_presently-available_methods, such as_the requirementxe2x80x2 for immunization of an animal and in_vivo steps would be very useful_particularly if it_made_it_possible to produce a_wide range of antibody types than_can be made using presently-available techniques_and if it made it possible to_produce human antibody types.
The present invention_relates to a method of_producing libraries_of genes encoding antigen-combining molecules_or antibodies; a_method of_producing antigen-combining molecules,_also reference to as antibodies, which_does not require_an in vivo procedure, as is_required_by presently-available methods;_a_method_of obtaining_antigen combining molecules_(antibodies)_of selected_or_defined specificity which does not require_an_in vivo procedure; vectors useful in the_present method and antibodies_produced_or obtained by the_method.
The present invention relates to_an in vitro process for synthesizing_DNA encoding_families of_antigen-combining molecules or proteins. In this process, DNA containing genes encoding_antigen-combining molecules is obtained and combined_with oligonucleotides which are_homologous to_regions_of the genes_which_are conserved._Sequence-specific gene amplification is then carried out_using the_DNA containing genes encoding antigen-combining proteins_as template and the_homologous_oligonucleotides as primers.
This invention also relates_to a method of creating diverse_libraries of DNAs encoding families_of_antigen-combining_proteins by cloning the product_of the in vitro process for_synthesizing DNA, described_in the preceding paragraph, into an appropriate_vector_(e.g., a plasmid, viral or retroviral_vector).
The_subject invention provides an alternative method_for_the_production of antigen-combining molecules, which_are useful affinity reagents for the_detection and neutralization of antigens and the delivery of_molecules to antigenic_sites. The claimed method differs_from_production_of polyclonal antibody molecules derived by immunization of live animals_and from_production of monoclonal_antibody molecules through the use of hybridoma cell lines in_that it does not require an in_vivo immunization step, as do presently available methods. Rather, diverse libraries_of genes which encode antigen-combining sites comprising a_significant proportion of an animal""s repertoire of_antibody combining