标准缓冲液的配制方法和PH计校正液(带目录).doc

标准缓冲液的配制及常用数据一、标准缓冲液pH值与温度对照表2二、常用缓冲溶液的配制方法21 甘氨酸盐酸缓冲液（0.05mol/L）22邻苯二甲酸盐酸缓冲液（0.05 mol/L）23磷酸氢二钠柠檬酸缓冲液24柠檬酸氢氧化钠-盐酸缓冲液35柠檬酸柠檬酸钠缓冲液（0.1 mol/L）36乙酸乙酸钠缓冲液（0.2 mol/L）37磷酸盐缓冲液38磷酸二氢钾氢氧化钠缓冲液（0.05M）49巴比妥钠-盐酸缓冲液（18）410Tris盐酸缓冲液（0.05M，25）51TE Buffer550TAE Buffer 配制方法：5马来酸 M=116511硼酸硼砂缓冲液（0.2M硼酸根）511硼酸-硼砂缓冲液（0.2M硼酸根）612甘氨酸氢氧化钠缓冲液（0.05M）613硼砂-氢氧化钠缓冲液（0.05M硼酸根）614碳酸钠-碳酸氢钠缓冲液（0.1M）615“PBS”缓冲液7三、实验室中常用酸碱的比重和浓度7四、硫酸铵饱和度的常用表8常见蛋白质分子量参考值 （单位：dalton）8常见蛋白质等电点参考值 （单位：pH）9元素的原子量表11PH计校正溶液配置的标准方法13一、标准缓冲液pH值与温度对照表温度四草酸氢钾0.05M邻苯二甲酸氢钾0.05M混合磷酸盐0.025M硼砂0.01M51.674.006.959.39101.674.006.029.33151.674.006.909.28201.684.006.889.23251.684.006.869.18301.684.016.859.14351.694.026.849.11二、常用缓冲溶液的配制方法1 甘氨酸盐酸缓冲液（0.05mol/L）X毫升0.2 mol/L甘氨酸+Y毫升0.2 mol/L HCI，再加水稀释至200毫升pHXYpHXY2.02.42.62.85050505044.032.424.216.83.03.23.43.65050505011.48.26.45.0甘氨酸分子量 = 75.07，0.2 mol/L甘氨酸溶液含15.01克/升。2邻苯二甲酸盐酸缓冲液（0.05 mol/L）X毫升0.2 mol/L邻苯二甲酸氢钾 + 0.2 mol/L HCl，再加水稀释到20毫升pH(20)XYpH(20)XY2.22.42.62.83.0555554.0703.9603.2952.6422.0223.23.43.63.855551.4700.9900.5970.263邻苯二甲酸氢钾分子量 = 204.23，0.2 mol/L邻苯二甲酸氢溶液含40.85克/升3磷酸氢二钠柠檬酸缓冲液pH0.2mol/L Na2HPO4（毫升）0.1mol/L柠檬酸（毫升）pH0.2mol/L Na2HPO4 （毫升）0.1mol/L柠檬酸（毫升）2.22.42.62.83.03.23.43.63.84.04.24.44.64.85.00.401.242.183.174.114.945.706.447.107.718.288.829.359.8610.3010.6018.7617.8216.8315.8915.0614.3013.5612.9012.2911.7211.1810.6510.149.705.25.45.65.86.06.26.46.66.87.07.27.47.67.88.010.7211.1511.6012.0912.6313.2213.8514.5515.4516.4717.3918.1718.7319.1519.459.288.858.407.917.376.786.155.454.553.532.611.831.270.850.55Na2HPO4分子量 = 14.98，0.2 mol/L溶液为28.40克/升。Na2HPO4-2H2O分子量 = 178.05，0.2 mol/L溶液含35.01克/升。C4H2O7H2O分子量 = 210.14，0.1 mol/L溶液为21.01克/升。4柠檬酸氢氧化钠-盐酸缓冲液pH钠离子浓度（mol/L）柠檬酸（克）C6H8O7H2O氢氧化钠(克)NaOH 97%盐酸（毫升）HCl（浓）最终体积（升）2.23.13.34.35.35.86.50.200.200.200.200.350.450.3821021021021024528526684838383144186156160116106456810512610101010101010 使用时可以每升中加入1克克酚，若最后pH值有变化，再用少量50%氢氧化钠溶液或浓盐酸调节，冰箱保存。5柠檬酸柠檬酸钠缓冲液（0.1 mol/L）pH0.1 mol/L柠檬酸（毫升）0.1 mol/L柠檬酸钠（毫升）pH0.1 mol/L柠檬酸（毫升）0.1 mol/L柠檬酸钠（毫升）3.03.23.43.63.84.04.24.44.64.818.617.216.014.914.013.112.311.410.39.21.42.84.05.16.06.97.78.69.710.85.05.25.45.65.86.06.26.46.68.27.36.45.54.73.82.82.01.411.812.713.614.515.316.217.218.018.6柠檬酸C6H8O7H2O：分子量210.14，0.1 mol/L溶液为21.01克/升。柠檬酸钠Na3 C6H5O72H2O：分子量294.12，0.1 mol/L溶液为29.41克/毫升。6乙酸乙酸钠缓冲液（0.2 mol/L）pH(18)0.2 mol/LNaAc(毫升)0.3 mol/L HAc(毫升)pH(18)0.2 mol/LNaAc(毫升)0.3 mol/LHAc(毫升)2.63.84.04.24.44.60.751.201.802.653.704.909.258.808.207.356.305.104.85.05.25.45.65.85.907.007.908.609.109.404.103.002.101.400.900.60Na2Ac3H2O分子量 = 136.09，0.2 mol/L溶液为27.22克/升。7磷酸盐缓冲液（1）磷酸氢二钠磷酸二氢钠缓冲液（0.2）pH0.2 mol/L Na2HPO4(毫升)0.3 mol/L NaH2PO4(毫升)pH0.2 mol/L Na2HPO4(毫升)0.3 mol/L NaH2PO4(毫升)5.85.96.06.16.26.36.46.56.66.76.86.98.010.012.315.018.522.526.531.537.543.549.555.092.090.087.785.081.577.573.568.562.5 56.551.045.07.07.17.27.37.47.57.67.77.87.98.061.067.072.077.081.084.087.089.591.593.094.739.033.028.023.019.016.013.010.58.57.05.3Na2HPO42H2O分子量 = 178.05，0.2 mol/L溶液为85.61克/升。Na2HPO42H2O分子量 = 358.22，0.2 mol/L溶液为71.64克/升。Na2HPO42H2O分子量 = 156.03，0.2 mol/L溶液为31.21克/升。（2）磷酸氢二钠磷酸二氢钾缓冲液（1/15 mol/L）pHmol/L/15Na2HPO4(毫升)mol/L/15KH2PO4(毫升)pHmol/L/15Na2HPO4(毫升)mol/L/15KH2PO4(毫升)4.925.295.916.246.476.646.816.980.100.501.002.003.004.005.006.009.909.509.008.007.006.005.004.007.177.387.738.048.348.678.187.008.009.009.509.759.9010.003.002.001.000.500.250.100Na2HPO42H2O分子量 = 178.05，1/15M溶液为11.876克/升。KH2PO4分子量 = 136.09，1/15M溶液为9.078克/升。8磷酸二氢钾氢氧化钠缓冲液（0.05M）X毫升0.2M K2PO4 + Y毫升0.2N NaOH加水稀释至29毫升 pH(20)X (毫升)Y（毫升）pH(20)X (毫升)Y（毫升）5.86.06.26.46.66.85555550.3720.5700.8601.2601.7802.3657.07.27.47.67.88.05555552.9633.5003.9504.2804.5204.6809巴比妥钠-盐酸缓冲液（18）pH0.04M巴比妥钠溶液0.2V盐酸pH0.04M巴比妥钠溶液(毫升)0.2N盐酸(毫升)6.87.07.27.47.67.88.08.210010010010010010010010018.417.816.715.313.411.479.397.218.48.68.89.09.29.49.61001001001001001001001005.213.822.521.651.130.700.35巴比妥钠盐分子量=206.18;0.04M溶液为8.25克/升10Tris盐酸缓冲液（0.05M，25）50毫升0.1M三羟甲基氨基甲烷（Tris）溶液与X毫升0.1N盐酸混匀后，加水稀释至100毫升。pHX(毫升)pHX(毫升)7.107.207.307.407.507.607.707.807.908.0045.744.743.442.040.338.536.634.532.029.28.108.208.308.408.508.608.708.808.9026.222.919.917.214.712.410.38.57.0三羟甲基氨基甲烷（Tris）HOCH2 CH2OH CHOCH2 NH2分子量=121.14;1M溶液为12.114克/升。Tris溶液可从空气中吸收二氧化碳，使用时注意将瓶盖严。1TE Buffer组成浓度：10mM Tris-HCl 1mM EDTA PH=8.0 配制量：500ml 配制方法：量取下列溶液于500ml烧杯中 1M Tris-HCl Buffer PH=8.0 5ml 0.5M EDTA PH=8.0 1ml 向烧杯中加入约400ml dd H2O均匀混合；将溶液定容到500ml后，高温高压灭菌；室温保存.50TAE Buffer 配制方法： 1。称量Tris 242g，Na2EDTA2H2O 37.2g于1L烧杯中； 2。向烧杯中加入约800ml去离子水，充分搅拌均匀； 3。加入57.1ml的冰乙酸，充分溶解； 4。用NaOH调pH至8.3，加去离子水定容至1L后，室温保存。 使用时稀释50倍（取20ml稀释至1L）或100倍 即1TAE Buffer 或 0.5TAE马来酸 M=11611硼酸硼砂缓冲液（0.2M硼酸根）pH0.05M硼砂（毫升）0.2M硼砂（毫升）pH0.05M硼砂（毫升）0.2M硼酸（毫升）7.47.67.88.01.01.52.03.09.08.58.07.08.28.48.79.03.54.56.08.06.55.54.02.0硼砂Na2B4O7H2O,分子量=381.43;0.05M溶液（=0.2M硼酸根）含19.07克/升。硼酸H2BO3,分子量=61.84,0.2M溶液为12.37克/升。硼砂易失去结晶水，必须在带塞的瓶中保存。11硼酸-硼砂缓冲液（0.2M硼酸根）pH0.05M硼砂（毫升）0.2M硼酸（毫升）pH0.05M硼砂（毫升）0.2M硼酸（毫升）7.47.67.88.01.01.52.03.09.08.58.07.08.28.48.79.03.54.56.08.06.55.54.02.0硼砂Na2B4O710H2O,分子量=381.43;0.05M溶液（=0.2M硼酸根）含19.07克/升。硼酸H2BO3，分子量=61.84, 0.2M溶液为12.37克/升。硼砂易失去结晶水，必须在带塞的瓶中保存。12甘氨酸氢氧化钠缓冲液（0.05M）X毫升0.2M甘氨酸+Y毫升0.2NnaOH加水稀释至200毫升pHXYpHXY8.68.89.09.29.450505050504.06.08.812.016.89.69.810.010.410.6505050505022.427.232.038.645.5甘氨酸分子量=75.07;0.2M溶液含15.01克/升。13硼砂-氢氧化钠缓冲液（0.05M硼酸根）X毫升0.05M硼砂+Y毫升0.2NNaOH加水稀释至200毫升pHXYpHXY9.39.49.65050506.011.023.09.810.010.150505034.043.046.0硼砂Na2B4O710H2O,分子量=381.43;0.05M溶液为19.07克/升。14碳酸钠-碳酸氢钠缓冲液（0.1M）Ca2+、Mg2+存在时不得使用pH0.1MNa2CO3(毫升)0.1MN2HCO3(毫升)20379.169.409.519.789.9010.1410.2810.5310.838.779.129.409.509.729.9010.0810.2810.57123456789987654321Na2CO210H2O分子量=286.2;0.1M溶液为28.62克/升。N2HCO3分子量=84.0;0.1M溶液为8.40克/升。15“PBS”缓冲液PH7.67.47.27.0H2ONaClNa2HPO4NaH2PO410008.52.20.110008.52.20.210008.52.20.3100ml8.5g2.2g0.4g三、实验室中常用酸碱的比重和浓度名称分子式分子量比重面分浓度%（W/W）当量浓度（粗略）N配1升1N溶液所需毫升数盐酸HCl36.471.1937.212.0841.1835.411.81.1020.06.0硫酸H2SO498.091.8495.636.0281.1824.86.0硝酸HNO263.021.4270.9816.0631.4065.314.51.2032.366.1冰乙酸CH3COOH60.051.0599.517.459乙酸CH3COOH98.06366.0磷酸H2PO435.051.7185.015,30,45(依反应而定)67(以15N计)氨水NH4OH0.9015670.90427.014.3700.9125.013.40.9610.05.6氢氧化钠溶液NaOH40.01.550.01953四、硫酸铵饱和度的常用表1调整硫酸铵溶液饱和度计算表（25）硫整硫酸铵终浓度,%饱和度10202530333540455055606570758090100每1升溶液加固体硫酸铵的克数硫酸铵初浓度，%饱和度05611414417619620924327731335139043047251656166276710578611813715018321625128832636540644949459269420295978911231551902252623003403824245206192530496193125158193230267307348390485583301930629412716219823527331435644954633124374107142177214252292333426522353163941291642002381783194115064031639713216820524528537546945326599134171210250339431503366101137176214302392553367103141179264353603469105143227314653470107190275703572153237753611519880771579079在25下，硫酸铵溶液由初浓度调到终浓度时，每升溶液所加固体硫酸铵的克数。2 调整硫酸铵溶液饱和度计算表（0）在0下，硫酸铵溶液由初浓度调到终浓度时，每100毫升溶液所加固体硫酸铵的克数。常见蛋白质分子量参考值 （单位：dalton）蛋白质分子量肌球蛋白myosin甲状腺球蛋白thyroglobulin-半乳糖苷酶-galactosidase副肌球蛋白paramyosin磷酸化酶aphosphorylase a血清白蛋白serum albuminL-氨基酸氧化酶L-amino acid oxidase地氧化氢酶catalase丙酮酸激活酶pyruvate kinase谷氨酸脱氢酶glutamate dehydrogenase亮氨酸氨肽酶glutamae dehydrogenase-球蛋白，H链-globulin, H chain延胡索酸酶（反丁烯二酸酶）fumarase卵白蛋白ovalbumin醇脱氢酶（肝）alcohol dehydrogenase (liver)烯醇酶enolase醛缩酶aldolase肌酸激酶creatine kinase胃蛋白酶原pepsinogenD-氨基酸氧化酶D-amino acid oxidase醇脱氢酶（酵母）alcohol dehydrogenase (yeast)甘油醛磷酸脱氢酶dlyceraldehyde phosphate dehydrogenase原肌球蛋白tropomyosin乳酸脱氢酶lactate dehydrgenase胃蛋白酶pepsin转磷酸核糖基酶phosphoribosyl transferase天冬氨酸氨甲酰转移酶，C链aspertate transcarbamylase, C chain羧肽酶 Acarboxypeptidase A碳酸酐酶carbonic anhydrase枯草杆菌蛋白酶subtilisin-球蛋白，L链-blobulin,L chain糜蛋白酶原（胰凝乳蛋白酶原）chymotrypsinogen胰蛋白酶trypsin木瓜蛋白酶（羧甲基）papain (carboxymethyl)-乳球蛋白-lactoglobulin烟草花叶病毒外壳蛋白（TWV外壳蛋白）TWV coat protein肌红蛋白myoglobin天门冬氨酸氨甲酰转移酶，R链aspartate transcarbamylase, R chain血红蛋白h(a)emoglobinQ外壳蛋白Q coat protein溶菌酶lysozymeR17外壳蛋白R17 coat protein核糖核酸酶ribonuclease或RNase细胞色素Ccytochrome C糜蛋白酶（胰凝乳蛋白酶）chymotrypsin220,000165,000130,000100,00094,00068,00063,00060,00057,00053,00053,00050,00049,00043,00041,00041,00040,00040,00040,00037,00037,00036,00036,00036,00035,00035,00034,00034,00029,00027,60023,50025,70023,30023,00018,40017,50017,20017,00015,50015,00014,30013,75013,70011,70011,000或13,000常见蛋白质等电点参考值 （单位：pH）蛋白质等电点鲑精蛋白salmine鲱精蛋白clupeine鲟精蛋白sturline胸腺组蛋白thymohistone珠蛋白（人）globin(human)卵白蛋白ovalbuin伴清蛋白conal bumin血清白蛋白serum albumin肌清蛋白myoal bumin肌浆蛋白myogen A-乳球蛋白-lactoglobulin卵黄蛋白livetin1球蛋白（人）1-globulin(human)2球蛋白（人）2-globulin(human)肌球蛋白Amyosin A原肌球蛋白myosin A铁传递蛋白siderophilin胎球蛋白fetuin血纤蛋白原fibrinogen-眼晶体蛋白-crystallin-眼晶体蛋白-crystallin花生球蛋白arachin伴花生球蛋白conarrachin角蛋白类keratins还原角蛋白keratein胶原蛋白collagen鱼胶ichthyocol白明胶gelatin-酷蛋白-casein-酷蛋白-casein-酷蛋白-casein-卵清粘蛋白-ovomucoid1-粘蛋白1-mucoprotein卵黄类粘蛋白vitellomucoid尿促性腺激素urinary gonadotropin溶菌酶lyso zyme肌红蛋白myoglobin血红蛋白（人）hemoglobin(human)血红蛋白（鸡）hemoglobin(hen)血红蛋白（马）hemoglobin(horse)血蓝蛋白hemerythrin蚯蚓血红蛋白chlorocruorin血绿蛋白chlorocruorin无脊椎血红蛋白erythrocruorins细胞色素Ccytochrome C视紫质rhodopsin促凝血酶原激酶thromboplastin1-脂蛋白1-lipoprotein1-脂蛋白1-lipoprotein-卵黄脂磷蛋白-lipovitellin芜菁黄花病毒turnip yellow vvirus牛痘病毒vaccinia virus生长激素somatotropin催乳激素prolactin胰岛素insulin胃蛋白酶pepsin糜蛋白酶（胰凝乳蛋白酶chymotrypsin牛血清白蛋白bovine serum albumin核糖核酸酶（牛胰）ribonuclease或Rnase(bovine pancreas)甲状腺球蛋白thyroglobulin胸腺核组蛋白thymonucleohistone12.112.111.7110.87.54.71; 4.596.8;7.14.7-4.93.56.35.1-5.34.8-5.05.8;6.67.3;8.25.2-5.55.15.93.4-3.55.5-5.84.86.05.13.93.7-5.04.6-4.76.5-6.84.8-5.24.7-5.04.0-4.14.55.8-6.03.83-4.411.8-2.75.53.2-3.311.0-11.26.997.077.236.924.6-6.45.64.3-4.54.6-6.29.8-10.14.47-4.575.25.55.45.93.755.36.855.735.351.0左右8.14.97.84.584左右元素的原子量表（录自1997年国际原子量表，并全部取四位有效数字）元素符号原子量原子序数锕银铝镅氩砷砹金硼钡铍铋锫溴碳钙镉铈锎氯锔钴铬铯铜镝镊锿铕氟铁镄钫镓钆锗氢氦铪汞钬碘铟铱钾氪镧锂镥铹钔镁锰钼氮钠铌钕氖镍锘镎氧锇磷镁铅钯钷钋镨铂钚镭铷铼铑氡钌硫锑钪硒硅钐锡锶钽铽锝碲钍钛铊铥铀钒钨氙钇镱锌锆AcAgAlAmArAsAtAuBBaBeBiBkBrCCaCdCeCfClCmCoCrCsCuDyErEsEuFFeFmFrGaGdGeHHeHfHgHoIInIrKKrLaLiLuLrMdMgMnMoNNaNbNdNeNiNoNpOOSPPaPbPdPmPoPrP