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ForanexplorationofthemoleculardimensionsofIDPs,coupledwiththoroughcontroloftheirmonomericstate,thecombinationofdynamiclightscattering(DLS)andstaticlightscattering(SLS)isamethodofchoice.IDPs：intrinsicallydisorderedproteins静态光散射：如果将溶液中的聚合物分子看作一个个各向同性的粒子,以一定频率的入射光照射这些粒子,它们不吸收入射光能量,而仅作为二次波源向各个方向发射与入射光频率相同的球面散射光,这种没有频率位移(即无能量变化)的散射称为弹性光散射(elasticlightscattering),也常称为经典光散射(classicallightscattering)、静态光散射(staticlightscattering)、time-averagedlightscattering在静态光散射情况下,通常用瑞利比来表征一个体系的散射能力和角度依赖性。当qRg1Aperceptibleangulardependenceofthescatteringintensity（可见的光强的角度依赖）canonlybeexpectedforparticleswithRG10nm.Thus,lightscatteringisnotanappropriatemethodformonitoringchangesofRGduringunfoldingandrefoldingofsmallmonomericproteins.Howeverasubstantialangulardependenceisobservedforlargeproteinaggregates.TheopticalconstantK=4π2n02(dn/dc)2/λ4NAdependsonlyonexperimentalparametersandthescatteringpropertiesofthemoleculesintheparticularsolvent,whichisreflectedbydn/dc.Theexactknowledgeofdn/dc,thedependenceofnonproteinconcentrationinthepresentcase,isveryimportantforabsolutemeasurementsofthemolecularmass.在一定温度下，测得不同角度时标准物、溶剂和溶液的散射光强I0（θ）、Is（θ）、I（θ），可得到相应得R（θ），实测或从高分子手册上查知n和dn/dC的数值，于是K成为已知的常数。KC/R（θ）是θ和C的函数。抽象地看，这是对应于一个空间曲面的函数z=f（x，y），其中x=q2，y=C，z=KC/R（θ）dn/dc可用微分折光仪测定，Measurementsoftherefractiveindexincrementdn/dcaremuchmoretime-consumingandrequiretheuseofdifferentialrefractometers,whicharealsofrequentlyusedasconcentrationdetectorsinchromatographytechniques.Therefore,instrumentsformeasuringdn/dccanbeobtainedfrommanufacturersoflightscatteringdevicesandchromatographysystemsaswell。However,itisrecommendedtotrytoobtainappropriatevaluesofdn/dcfromtheliteratureforparticularbufferandsolventsbeforestartingtediousexperiments.Therefractiveindexnanddn/dcweremeasuredusinganAbberefractometer(KarlZeiss,Jena)ThestandardreferencesampleforcalibrationofanSLSinstrumentistoluene(甲苯，ultrapure,e.g.,MerckUvasol).Tolueneshouldbefilledintoacarefullycleanedsamplecell.热力学参数---第二维利系数A2与huggins参数χ1一样，表征高分子“链段”与溶剂分子之间的相互作用。A2可用于表征稀溶液中大分子（如蛋白质）间相互作用强度。Thesecondosmoticvirialcoefficient,B22,isausefulparameterincharacterizingprotein–proteininteractionsindilutesolutions。Inproteinsolutions,B22isrepresentativeofinteractionsbetweentwoproteinmolecules.ApositivevalueofB22isindicativeofrepulsiveinteractionsandanegativevaluerepresentsattractiveinteractions.Aproteinformulationwithnetrepulsiveinteractionsispreferableasattractiveinteractionscanleadtoself-associationandaggregationofproteins(LeBrunetal.,2009,Saitoetal.,2011andSalujaetal.,2007b).Analyticalultracentrifugation(AUC)andstaticlightscattering(SLS)areroutinelyusedtechniquestodeterminethisparameter(GeorgeandWilson,1994,Liuetal.,2005andMuscholandRosenberger,1995).However,ithasbeenarguedthattheparameterobtainedfromstaticlightscatteringisB2notB22becauseinSLS,protein-cosoluteinteractionsalsocontributetothesecondvirialcoefficient(Winzoretal.,2007).Nonetheless,thereareseveralstudiesthatshowtheuseofSLSinunderstandingpropertiessuchasproteinsolubility,self-associationofproteins,andcrystallization(GeorgeandWilson,1994,Salujaetal.,2007aandSchereretal.,2010).Staticlightscattering(SLS)iscapableofmeasuringmolarmasseswithintherange103–108g/molandisthereforeidealfordeterminingthestateofassociationofproteinsinsolution.wherethepolymerglobulesareconsiderablysmallerthanthewavelengthoftheincidentlight,P（θ）willreduceto1(Rayleighapproximation).Thisconditionmaybeverifiedbydeterminingthediameteroftheglobulesbydynamiclightscattering.当颗粒直径小于100nm时，由He-Ne激光器产生的散射强度的角度依赖性可以忽略，无需完整的Zimm作图来求A2，只需在一个角度做一系列浓度的测定即可。可通过瑞利公式计算。Debye作图法。NanoZS型检测仪可以测定水力学直径Dh、zeta电位和分子量，并在检测分子量同时提供第二维里系数A2（Thesecondosmoticvirialcoefficient）。动态光散射：多普勒效应：当单一频率（～1015Hz）的入射光被散射时，如果粒子处于静止状态，那么散射光的频率将会同入射光的频率相同，即弹性散射。实际上，由于粒子的无规则brown运动，散射光的频率将会随着粒子朝向或背向监测器的运动面出现极微小（～105～7Hz）的增加或减少，使得散射光的频谱变宽：当产生散射光的分子朝向监测器运动时，相当于把散射的光子往监测器送了一段距离，使光子较分子静止时产生的散射光更早到达监测器，也就是散射光的频率增高了，反之，背向运动则使频率降低。显然，频率变宽的幅度（线宽）是同粒子运动的快慢联系在一起的。如果测得线宽，就可以得到粒子运动快慢的信息，然后粒子的brown运动实在太慢，所引起的Dopplar效应（即频率变宽）仅有一亿分之一左右，可利用快速光子相关仪在时间空间中通过时间相关函数G（t）来测得。Whenparticlemotionisnotrandom(e.g.sedimentation,creaming)DLSisnotthecorrecttechniquetouseRH=KDT/6πηDLikecalculationsofdn/dc，viscositymeasurementsmaybecomemoretime-consumingthanatypicalSLS/DLSexperimentitself.Therefore,itisusefultochecktheliteratureforappropriatedata。DLSanalyzestheabove-mentionedtemporalfluctuationsoftheinstantaneousintensityofscatteredlight.DM表征与真实颗粒具有相同质量的硬球直径，DR表征与真实颗粒具有相同几何中心的球体直径，回转直径Dg与分子中原子质量和原子距离分子质量中心的距离有关，DH描述溶液中和真实颗粒具有相同扩散速率的假想硬球的直径。由于大分子在溶液中以非球形、动态的、溶剂化的形式存在，所以从颗粒扩散性能计算出的直径是动力学的、水合的、溶剂化颗粒的假想大小，称为水力学直径。水力学直径与颗粒本身性质（如形态、分子量）相关，还受周围溶液环境因素的影响，如Ph、离子强度、溶液粘度、吸附性物质等pH：一般处于等电点附近时颗粒的水力学直径最小，当pH大于或小于等电点时，颗粒水力学直径增大。离子强度：蛋白质水力学直径随离子强度增大而减小。表面活性剂：对水力学直径的影响可以通过表面活性剂在带电颗粒表面的吸附来解释。开始表面活性剂浓度增加，水力学直径增大，当表面活性剂在微粒表面形成稳定的胶束层，吸附达到饱和，微粒与表面活性剂复合物水力学直径达到最大。DLSinstrumentsdonotrequirefurthercalibrationaslongasallexperimentalparametersusedfordatatreatment(e.g.,l,y,n,T,_)areproperlydetermined.However,testexperimentswithsolutionsofmonodisperselatexspherescouldbeusedtoverifytheproperfu