PROTEIN PRODUCTION - Bio-Rad
Contents
1.2.
3.
4. amp
5. gt EL El es
6. JOA SUOHBIOBHOI le HBIDBHOI C Interfacial Refolding Caio PO4 OH
7. Bi O W 2 A QO 9 O 33 Rosalind Kim PhD 4 livr Rosalind Kim US National Institute of General Medical Science NIGMS Protein Structure Institute 9 Berkeley Structural Genomics Center BSGC
8. Biotechnolgy and Bioengineering Marcel Dekker Dr Maurits Dekker fax e mail internet 1988 Ion Exchange Chromatography of Proteins
9. 0 User s Voice 0 TransFectin F9 TransFectin 50 9 H primary cultured cortical neuron U SiLentFect
10. siRNA in situ hybridization
11. ELISA Explorer 3
12. 3
13. 1979
14. IntervieW 1 4
15. 2006 1 Expression Proteomics Tools for Protein Separation and Analysis 4 gt J ReadyPrep Aurum 5 ReadyPrep
16. hr RT LB amp 2 No PR PO
17. coven Story Amino residueSs 4 Classical cation exchande Elute with neutral salts NaCl or PO4 1 P site C site 1mM C site 1 50 500 mM Carboxyl clusters Classical chelation modulated by ion exclusion 15 60x stronger tnan ionic Interactions alone Will not elute at
18. Berkeley Structural Genomics Center http www strgen or9 Dr Kim s group at the BSGC http www strgen org aboutcloning html ProteoCell Biotechnologies Inc http proteocell com Genitope Corporation http www genitope com The NIGMS Protein Structure Initiative http www nigmS nih goV psi The Protein Data Bank the single worldwide repository for the processing and distribution of 3 D biological macromolecular structure data http www rcsD org pdb Miguel Retamanl Chamow
19. Y DCode DCode DCode E HHHMOCUS Interview
20. 9 IOA SUOnBID6HO J HHHMOCUS Concepf 4 Society and Education PCR DGGE Denaturing Gradient Gel Electrophoresis
21. eyond 3 2005 10
22. 03 5811 6290 GY 03 5811 6697 0O UU W 2 A QO am 9 O oofT 2005 11 15 17 2005 11 23 25 CE 2005 12 7 10 4 PCR RNAi RNAi PCR RNAi
23. Retamal Genitope Corporation NHL
24. hepatic stellate cell HSC HSC TNFz TGF interferon y IFN y HSC IFN y IFN y TACE TNF converting enzyme HSC TNF
25. RUBY Flamingo Flamingo RUBY 120 excitation minus protein 100 excitation plus protein emission minus protein 80 emission plus protein 5 _ c 60 We 40 LL 20 250 350 450 550 650 nm 1 Flamingo 50 mM
26. FDA 10 3
27. DGGE DCode 6 User s Voice DCode DCode DGGE A
28. 9 Prortein Fingerprinting
29. Retamal 100 Retamal ProteoCell Biotechnologies ProteoCell Biotech nologies ProteoCell Biotechnologies current Good Cl
30. WM 1 GFP 4 8 PV92 PCR Informadcs 8 SPP DNA
31. 2 3 Experion Caliper Life Sciences LabChip RNA Experion 10 260kD Experion Pro260 Experion Pro260 10 30
32. gt SD HIV AIDS HIV PA HIV 1 2 HIV 1 2 PA HIV HIV PA HIV 1 2 e 1 HIV 1 2 PA SFD HIV 1 PA HIV Ag Ab 1
33. 0 1 mg ml 0 4 mg m1l Experion Pro260 Experion Pro260 10 3 2 8 Pro260 2 1 4 1 95 5 84 1 Pro260
34. Native 2 5 ml g mg pH 10 200 250 1 pH pH pH pH
35. 9 IOA SUOnBIDGHO i OT Bridde PV92 PCR Informatics nr WWW a rm It aC me ng 16 SSH
36. Data Analysis 3 D Structure and Function 3 Data Analysis 3 D Structure and Function PAGE Kim PSI
37. PCR MiniOpticon PCR Benchmark ELISA PCRH MJ Mini MiniOpticon PCR MJ Mini 48 MJ Research MiniOpticon PCH
38. CHT 1 CHT 2 3 4 CHT 5 Introduction about Bio Rad s industrial scale chromatography columns and cleaning in place capabilities France Bio Rad Laboratories Process Hardware Research amp Development Manager Pierre Vidalinc 6 Purification of monoclonal IgG with CHT ceramic hydroxyapatite USA Bio Rad Laboratories R amp D manager of Process Applications Dr Peter Gagnon 7
39. product0OGO 600 cm retention time 150 cm 20 3 21 8 mg ml IMAC Profinity His tag 6 IMAC Profinity A 1 B His tag IMAC His tag
40. 1 3 Aurum IgG IgG a Visit us On the Web at discover Dbio rad cO jD 116 0014 5 18 TEL 03 5811 6270 FAX 03 58 1 1 6 7 Helping Science Serve Mankind whatS 1 10
41. Rosalind Kim Kim 10 3 NIGMS Protein Structure Initiative PSI 5 Kim PSI BSGC Component Project Leader Cloning Expression and Purification Berkeley Structural Genomics Center Berkeley CA USA PSI
42. IMAC 20mM 50 v v 20mM 20cm 43 psi 3 bar producte GO A Profinity IMAC 50 50 40 40 30 30 8 gO A B Resin 1 from supplier A eg 0 C Resin 1 from supplier B 50 dD 3 eg0 600 800 1000 0 200 400 B00 800 1 0OO Flow rate crm hr 8 Maximum Flow Rate at 43 psi cm hr Viscosity cP Profinity IMAC Supplier A Resin 1 Supplier B Resin 20 mM 1 06B 00 d70 750 1 MNaOH 1 623 780 828 560 amp 6M 1 821 610 590 370 8M 2 062 550 530 250 1 IMAC 100 200 cm h O 2 AC D
43. g TransFectin 10
44. pH IMAC Profinity His tag His tag IMAC CO O1 CO Ei ET a 2 kD i a Pe B Resin Yield Purity Profinity IMAC 4 3 mg 93 Resin 2 supplier A 6 3 mg 8196 Resin 1 supplier A 4 0 mg 7896 SuDDlier g 9 mg 8996 SuDDlier C 2 8 mg 92 7 Ni IMAC 32 kD His tag His tag Anabaena
45. 1 Marcel Dekker 3 Taylor amp Francis Rusell Dekker Dekker 1930 2 J Polymer Science vol 26 xv xvi 1988 Kirk Othmer Encyclopedia 2 lon Exchange Chromatography of Proteins Dekker 1998 10 Taylor amp Francis 2008 20 J C Giddings Dynamics of Chromatography 1965
46. N Signal Transduction 2
47. 1
48. Caco 2 siLentFect siRNA MLCK1 Tech Note 5370 siRNA siRNA Tech Note 5399 Web http discover bio rad co jp TransFectin siLentFect Tech Note 5343 Gene Expression
49. lt 76 6 2 4 3 2 1 4
50. pI
51. Molecular Weightt MW Experion MVW SDS PAGE R Pro260 CV 1 05 SDS PAGE CV R Excel 3 78 1 Rr MW Pro260 8 56 8 49
52. Bio Plex tie et COOH G ee Bio Plex R amp D
53. 20 10 IEF Native 29 5X18 8X16 cm 2 85 kg Native MALDI TOF MS LC MS MS 2 Ordering Inforrmation 170 2800 490 000 PharosFX PharosFX Plus PharosFX 3
54. IMAC Profinity His tag tag IMAC 7 250 kD 8 IMAC Profinity His tag Ni IMAC Profinity His tag eIMAC Profinity 200 600 cm retention time
55. Kim Miguel Retamal ProteoCell Ciotechnologes Miguel Retamal 11 Retamal
56. Target Selection Characterization Functions and Interactions Cloning amp Expression His tag Purification IMAC Characterization X Ray Data Collection X X X 3 Crystallization X Ray Data Collection Function amp Interaction 3
57. His tag IMAC Profinity UNO sphere IDA 60um 1 His tag Materials Properties e Anabaena sp PCC7120 32 kD His tag e His GFP e 250 kD His tag e 75kD His tag
58. 10 COSFectin COS 7 COS 1 COS 3 HEKFectin a HEK 293 293H 293F 293T 293Phoenix Bosc 0 5 0 5 1 0 1 5 2 0 3 0 4 0 HEKFeoHn 20 acompetrB A es 3 0 05 S 0 5 1 0 1 5 2 0 3 0 4 0 Amount of transfection reagent ul well 1 Q pCMV SPORT 8 gal 24 COS HEK293 Ordering Inforrmation COSFectin Cell Specific Lipid 0 5 ml 4C 170 3370 32 000 COSFectin Cell Specific Lipi
59. 2 CA Sigma Aldrich Inc BSA National Institute of Standards and Technology NIST Boehringer Mannheim E coli 163 2110 53 kD PBS E coli 2 mg ml BSA 200 g ml 2 5 2 000 ng 1 CA 9 8 b BSA
60. 1996 Ig NHL B B Ig Genitope Trg IgG Ig Ig Ig B
61. 500 psi 2 3 IMAC Profinity IMAC Profinity 1 ml HR5 5 201 4 75kD His tag 1 ml
62. Helping Science Serve Mankind HR_HP bio rad com
63. References OS of liquid chromatography 9 16 Chromatography Vol 25 No Supplement 2 39 40 2004 10 29 Vol 2005 225 2005 09 25 Absorbance at 280nm Absorbance at 280nm Absorbance at 280nm Absorbance at 280nm Cp 0 2 50 40 30 0 1 20 10 0 0 0 5 10 15 20 Time min 2 C CHT C Native 0 2 50 Native DT 40 30 0 1 20 10 0 5 10 15 20 Time min 3 C CHT C C Native a 0 9 10 19 20 Time min 4 C
64. 3 4 4 9 IOA SUOnBIDHO DNA
65. T 9 IOA SUOnBID6HO Our FOCUS Concept 2 Protein Separation CHT 3 CHT P42 RW CHT CHT i a lg lt
66. 8
67. Bio Plex TNF Bio Plex HSC class I IFN y Bio Plex
68. Genitope Corporation Process Science Chamow Chamow Genitope Corporation lg 75
69. eFlamingo 0 10 20 30 40 50 60 ng B galactosidase 4 Flamingo SDS PAGE Criterion 4 20 Tris HCI Flamingo Molecular Imager FX Quantity One A 0 25 4ng BB 4 60 ng C 60 960 ng B RUBY i fF Fi a Es se CE Ep H EE LL T ee
70. 20 mM 1M NaC1 1M NaOH 6 M gt 8 M gt 20 gt 70 IMAC Profinity pH 4 0 1N NaOH 4 IMAC Profinity 5 Profinity IMAC 15 4 mgml SUuDDIEr A Bsin 1 2 5 Supplier 14 1 mmml 2 0 15 1 0 4 rg 0 5 0 0 10 20 30 Run tirme Min 6 IMAC 10 His tag
71. SDS PAGE 20 10 1 10 MW 1 1 4 20 SDS 2 PAGE Experion Pro260 SDS PAGE SE N Expedon _ SpS PAGE_ Pro260 Protein of Conc of _ Sample Wells ng SD Reproducibility Lanes Density SD Reproducibility BSA gg 10900 4 70 4 30 12 1 11 0 09 8 4196 A s 8 CA 30 161 00 700 11 009 12 0 15 005 31 3796 SDS PAGE ProgE lacdder i 10 29 157 10 21 00 13 0Q9 16 0 40 0 03 7 44 20 29 182
72. Quantity One CA 2 Pro260 rr7 0 97 0 99 n 5 4 20 Tris HC1 r 0 96 0 97 n 4 Experion r 0 98 CA 10 1 000 ng 1 Experion Experion SDS PAGE Experion Experion
73. HP DNA Gene in a bottle PCR PV92 PCR ALDH2 Informatics PCR ALDH2 2 DNA
74. Protein A DNA CHT ProteinA DNA EC CHT a CHT 98 Web DNA es http www bio rad com ad CHT Visit us on the Web at discover bio rad co jp 116 0014 5 18 TEL 03 5811 6270 FAX 03 58 1 1 6 7 Helping Science Serve Mankind Dr Je
75. 260 kD Upper SDS PAGE SDS PAGE Criterion Tris HC1 4 20 T Criterion 4 20 T Experion Pro260 5 4 1 2xLaemmli 4 1 95C 5 200V 55 Bio Safe CBBG 250 1 GS 800 Quantity One 1 D 3 three replicates 4 n 12 CA
76. SSC1I TI DNA SSCI Biotechnology Explorer Gene in a bottle DNA TpGLO
77. 10 50 mg ml 5 50 mb ml 10 20 40 80 m 4 lt CHT Type 1 Type I gt or 6 1 RN 10 um 4X100mm Pressure 23 bar A A 1 mM ph6 8 oum ff ODM 9 40 um S Pressure 2 bar 80 um AA 0 5 bar 4 8 12 4 8 12 LU 8 12 min Time min Time min Time min s ait tad FiaFO4 Ce mi
78. CHT C C CHT C C C CHT C Interfacial Refolding
79. Retamal Steven Chamow PhD Vice President of Process Sciences Genitope Corporation Redwood City CA USA ha i Steven Chamow NHL
80. URL http Shu chem yamaguchi u aC jD 10 T 6 Diffusion 9 IOA suUonelpedolg 1 HHHMOCUS ConceDpf 3 Signal Transduction
81. 20 ProteoCell Biotechnologies BSGC Chamow Steven Chamow
82. 1 30 COOH 100 1ml 1 10 1 6 150kD BSA Tris PBS pH 7 4 2 EDC 1 ethyl 3 3 dimethylaminopropyll carbodiimide hydrochloride S NHS N hydroxysulfosuccinimide 1 4C 1 3 Bio Plex
83. 2 SDS
84. 2 FDA Chamow
85. High Efficiency Low Toxicity High Efficiency 24 well platee70 confluencee Serum Relative Activity 4 ci 0 SiRNA siRNA OOS7 He 299 a RE IransFectin Competitor L2 Competitor LP _ Competitor F siRNA TransFectin i i siLentFect Lipid Reagent siRNA siRNA siRNA siLentFect
86. TEFF Tangential Flow Filtration ProteoCell Biotechnologies His Lysis IMAC Miguel Retamal UO IOA SUOIHBIODBHOI ew hepedolg Kim
87. 4 2 Protein A purified human IgG1 CHT type I 20 micron pH 6 5 phosphate NaCl grad gradient plan A plan B 5 3 5 NaC1 Purity CHT Aggregates DNA Endotoxin A CHT
88. MW A Experion i 0 500 1 000 1 500 gUOO B SDS PAGE gt i 3 CC 1 T3 0 500 1 000 1 500 2 000 Expected GA ngrul 2 Experion Pro260 SDS PAGE CA Pro260 A SDS PAGE B r 0 9887 A 0 9626 B g 6 UJ O O lt O 23 product GOGOS 1 Experion Pro260 n 25 SDS PAGE n 25 MW SD
89. ABO ELISA Gene in a Bottle DNA DNA DNA DNA DNA DNA
90. 200 cm h 2 43 psi 3 bar 10 3 Fu TIB min 50 mM EDTA 50 mM 300 mM NacCI pH 7 5 50 mM 300 mM NaCI pH 4 0 100 mM NiSO4 pH 4 0 50 mM 300 mM NaCI pH 8 0 1 0N NaOH 4 BioLogic DuoFlow IMAC Profinity 1 ml 201 1 50 75 kD His tag 3 51mg 250 Results and Conclusion IMAC 43 bar 20 cm IMAC Profinity 10 mm 5 A
91. 5299 product GOGOS IMAC Profinity His tag Danni Wang Xuemel He Shane Petersen Fang Fand Wu Yueping Xu Tanis Correa and Hong Chen 6000 James Watson Drive Bio Rad LaDoratorles Inc Hercules CA 94547 USA Method IMAC His tag IMAC IDA NTA Cu Ni Co Ca Zn IMAC
92. Emission 535 nm 10X 0 O rderind Inforrmation 10XEF amingo 20ml 200ml 4C 161 0490 7 000 10XFlamingo 100ml 1L 4C 161 0491 23 000 10XFlamingo 500ml HL 4C 161 0492 99 000 2 whats NEW PDQuest 8 0 Advanced amp Basic PDQuest 2 Version8 0 2
93. 6 12 kg 600 Kim 10 20 mg co77 His E coli IMAC His SDS SDS BSGC Target Selection M pneunomiae
94. PharosFX amp 1 532 nm 2 488 nm 532 nm 3 488 nm 532 nm 635 nm 3 PharosFX Plus RI 1 5 cm
95. 0O UU W 2 A QO 2 9 HHHMOCUS
96. DGGE 16S rRNA DGGE DGGE
97. O rderind Inforrmation PharosFX PharosFX Plus whats NEW IOA SUOIHBIDBO HPIDPHO whatS Flamingo Flamingo 1 Flamingo 0 25 0 5ng UV CCD Excitation protein Excitation protein Emission protein
98. PCR DGGE DGGE DNA GC GC PCR 2 DNA CC 2 GC Y Y DNA DNA 16s rDNA TT ETI I GC 2 CN DGGE
99. PDQuest Ver8 0 Advanced Basic 2 Basic Advanced Basic 3 PDQuest Advanced PDQuest Ver8 0 03 5811 6271 O rderind Inforrmation PDQuest 8 0 Advanced Software PDQuest 8 0 Basic Software TTP 4 Hed hmapwbsegewrd rm iR Digiig ekers Ja Ra Se titani RE
100. RNA interference RNAi mRNA 2 RNA mRNA gene silencing 1998 mRNA 20 25 2 RNA siRNA gene silencing siRNA PCR siRNA 2 5 kb mRNA 3 2 10
101. 1 His tag 1 1 1X30 cm Amicon Bio Scale MT2 GE Healthcare HR 5 5 IMAC Micro Bio Spin 1 000 g IMAC His tag SBS Medel DV 100 Stony Brook Scientific Ltd
102. 37 KDa 37 37 23 35 9 1 37 10 8495 15 8196 5 33 59 31 0 E B D RT B 38 G 95 d 1896 Experion 25 KDa 25 24 91 29 0 F 6 1H 19 Ur 1 B 36 1 2 d795 0 17 1 1 A B C D E F 2 Experion Pro260 4 20 Tris HC1 M ee 5 a O E coli g JJ o o N _F EL EE E Pro260 35 ls O o 3 Quantity One
103. 364 8501 6 100 TEL 048 593 1236 mailto kobayashi n kitasato or jp 3
104. CHT 70 CHT CHT Geltec 7 4
105. Experion SDS PAGE Expected MW kD MW kD Accuracy Reproducibility MW kD AcCCuraCy Reproducibility 14 3 14 23 0 12 0 4996 0 82 12 10 0 42 15 38 3 47 18 4 18 82 0 13 2 269 0 6996 14 70 0 50 20 1196 3 4096 26 6 26 10 0 27 1 8696 1 05 24 00 0 69 9 77 2 88 36 5 33 37 0 29 8 569 0 86 32 20 1 00 11 7896 3 119 45 44 43 0 34 1 26 0 77 42 30 1 60 5 00 3 7896 55 56 32 0 47 2 3996 0 84 51 40 1 37 6 5596 2 6796 66 71 60 0 70 8 4996 0 97 66 70 1 78 1 069 2 679 6 b 97 95 44 0 67 1 61 0 7196 96 20 2 75 0 8296 2 8696 116 123 00 0 55 6 02 0 4596 110 40 3 06 4 8396 2 7796 MW CV Upper X100 260 kDa Lower 10 kDa CV CV
106. 4X35 mm Type I 10 gm ph6 8 10 mM 400 mM gt 2 microglobulin ee ses Pg e PC e 09 4 ee PO 4 TOT Py 4 Pd Pr Pr ee Pr PC 9 4 ATT PO 9 PC PP ee PT PC Pr Pr Pr Pa Pr Pr Pr Ya Pr PL PC Pr oe oe tHe e
107. Experion Pro260 4 1 2 3 4 SDS PAGE Experion SDS_PAGE Experion Pro260 SDS PAGE E coli 2 D
108. Innovation 30 RNA RNA 1 l 4 F a Web http www bio rad com ad experion Visit us On the Web at discover Dbio rad cO jD 116 0014 5 18 TEL 03 581 1 6270 FAX 03 58 1 1 62 Helping Science Serve Mankind Z1210 0604A
109. C 2M C C C C 1 c 2M 1 d 2M 1 e 5 2M C 100 C
110. PV92 PCR Informatics PV92 PCR Informatics PC DNA PCR PV92 Alu Alu
111. PartI Giddings Part 30 3 Marcel Dekker Separation Science and Technology J C Giddings 2 aggressive Steven Cramer Editorial board member Synergen lt
112. 3 4 A B D E 3 4 F Experion Pro260 4 20 Tris HC1 SDS PAGE SDS PAGE Experion Pro260 10 30 Experion Pro260
113. 3 siRNA 1 12 uM 26 2 0 3 21 6 1 9 19 8 0 4 11 7 1 6 1 Saxena Jonsson Z0 Dutta A Small RNAs with imperfect match to endogenous mRNA repress translation Implications for off target activity of small inhibitory RNA in mammalian cells J Biol Chem 2003 Nov 7 278 45 44312 9 2 Du Q Thonberg H Wang J Wahlestedt C Liang Z A systematic analysis of the silencing effects of an active siRNA at all single nucleotide mismatched target sites Nucleic Acids Res 2005 Mar 21 33 5 1671 7 3 Yoshida K 0kamura H Amorim BR 0zaki A Tanaka H Morimoto H Haneji T Double stranded RNA dependent protein kinase is reduired for bone calcification in MC3T3 E1 cells in vitro Exp Cell Res 2005 Nov 15 311 1 117 25 0O W 2 A QO 9 2 PhD IFA ELISA 1 PC 10 PhD Qg 0 I O O 5
114. 50 mM 0 3 M NaCI 8 M pH 8 0 IMAC Micro Bio Spin His tag 250 mM A 3g SDS PAGE 1 8 10 Precision Plus Protein 2 3 A 2 4 A 1 5 IMAC Profinity 6 B 7 C B His tag Profniy IMAC Supplier A rsin 3 Supplier 10 15 3 B Run volume ml 1 2 8383 4 5 tman a E250OkD a C a Resin Yield Purty Profinity IMAC 3 6mg 86 SupplierA 2 1mg 3696 Supplier 1 1 mg 20 8 IMAC 250kD His tag A CV 5
115. gt ee 0 i SC laren PO mm NNO EF NaCI 15 60 RN CHT Type I Type II 2 1 Type 800 1 000 A Type a
116. 3i PE ia Ea 13 DNA 2 13 3 3 1 40 1
117. Experion Pro260 MW 10 R R 10 kD R 1 2 260kD R 2 0 25kD Rs 1 5 10 Pro260 MW Pro260 SDS PAGE MW 1 13
118. 12 2 siRNA 40 nM 3 RT PCR iCycler iQ MyiQ iQ SYBR Green Supermix Sequi Blot PVDF ChemiDoc XRS J 1 A B siRNA 909 RT PCR 8 siRNA 40nM 10 c 05 8 cont A B siRNA 40nM 2 RT PCR siRNA cont siRNA A B RT PCR A A B
119. 2 siRNA siRNA A B RT PCR mRNA A A B A mRNA B mRNA siRNA B siRNA siRNA mRNA RNA siRA siRNA
120. Genitope Genitope Genitope 1 Ig Genitope zIg Chamow
121. 5 25 60 65 100 S N product0OGO A ExDerion del View B SDS PAGE C Experion electropherogram 2 5 ng ul CA CA ng EA ng Ph Ph lt a 3 Ro qr MT NT AV Y MT Mr kD MVWi kL i la las lm boos i FE 260 ea 150 ws 100 ws 75 1CMD 75 50 m rr _ I f 20 20 1 Experion SDS PAGE CA CA Experion Pro260 A 4 20 Tris HCI SDS PAGE B C Experion 2 5 ng L CA 10 ng CA 1 2 kD Lower
122. DNA GC Y DNA DGGE DGGE DGGE DCode F DGGE
123. IMAC Profinity 43 bar A 1 B IMAC 2 IMAC Profinity 2 400cm 100 psi A B IMAC A 1 B IMAC 50 psi IMAC Profinity 43 100 psi IMAC Profinity 3
124. 116 0014 5 18 TEL 03 5811 6270 FAX 03 58 1 1 6 7 Helping Science Serve Mankind 7777777 gt SiRNA mRNA siRNA RNA mRNA siRNA siRNA RNA siRNA siRNA
125. SDS SDS SDS C C a b C d 6 1 C a b 2M c d 2M e 2M Urea C C 5 mg ml 2 M
126. 4 Experion Pro260 SDS PAGE M O 33 4 0 1 0 3 R R 1 17X ts ti wi ws t 1 t 2 w1 1 ws 2 R R 1 5
127. CA 3 E coli A Experion Pro260 Upper B SDS PAGE Quantity One product0OGO 3 Experion Pro260 SDS PAGE Observed MW kD Observed Difference Expected MW kD Experion SDS PAGE Expected Difference Experion SDS PAGE A Experion 100 KDa 100 100 11 97 2 9 1295 5 5495 b STM 5 15 9 1 B Experion 75 KDa YO 4 18 BU 1595 1B 51 BSA B 1 BD nr C 5g 50U na B U95 195 op Experion 50 KDa 50 50 09 50 8 D Experion
128. NN o Sa DCode Society and Education 3 1 ribosomal protection protein RPP RPP lt RPP 2
129. pH 4 0 100g ml 165nM Flamingo 8M 10 mg ml 100 480 nm 610 nm 0 go DP PD NH SH Hoo bh YN OP or So ng A myosin gal sh BSA Ovalb carb anh tryp inhib IlySOZ aprot 532 nm laser excitation Flamingo 100 g ml 1 Flamingo 4 80 g ml 2 Elamingo 532 nm 0 25 ng 300 nm UV 0 5 ng 3 400 350 300 250 200 BSA
130. 1 3 lechnicaol Heports mRNA RNA siRNA 2 siRNA mRNA RNA RNA PKR double stranded RNA dependent protein kinase PKR eIF2z eukaryotc initiation factor 2 alpha mR
131. 15L 370L 1992 1993 Yamamoto Model 1 1 Martin 30 The chromatographer should beware of becomng so in love with his technique that he neglects other methods of separation Other methods are usually more adaptable when large quantities are in question A J P Martn Nobel Laureate 1
132. events BIO RAD discover bio rad co jp Exhibit Dates Location 3 79 2006 3 8 10 CD http jps79 umin jp Proof of concept Bio Plex 2006 3 8 5F C 12 00 13 00 Exhibit Dates Location 5 EXPO 2006 5 17 19 http Awww bio expo jp Exhibit Dates Location 20 2006 6 18 23 20th IUBMB International Congress of Biochemistry and Molecular Biology and 11th FAOBMB Congress http vww congre cOo jp iubmb indexj html 2006 07 2006 07
133. 02 6 1014 1822 26 30 34 38 42 46 50 Cyo factor VI 1 0001 1001 02 6 10 14 18 9 26 30 34 38 42 46 50 Cycle iQ5 PCR 4 DNA 10 0 5ng 50 pg 50 1 iQ Multiplex Powermix e Tubulin FAM 96 4 r 0 998 GAPDH HEX 94 9 0 999 IL 1 Texas Red 101 8 P 0 999 FactorVll Cy5 101 6 r 0 997 O rderind Inforrmation PCR iQ Multplex Powermix 50 Lo 170 8848 28 000 iQ Multiplex Powermix 200 170 8849 88 000 whatS NEW COSFectin Cell Specific Lipid HEKFectin Cell Specific Lipid COSFectin HEKEFectin Cell Specific Lipid COS COSFectin HEK 293 Ti ep TransFectin
134. 2005 12 13 15 3 Bio Plex Bio Plex Bio Plex
135. 5 6 10 Q10 10 His tag IMAC Profinity IMAC UV SAS PAGE SDS PAGE Criterion Tris HC1 Quantity One 7 8 200 Profinity IMAC SUDDMier A reSin 1 Supplier B Cn Pressure psi 0 400 800 1 200 1 600 2 000
136. CHT C C Native 0 0 5 10 15 20 Time min 5 C CHT C C Conductivity mS cm Conductivity mS cm Conductivity mS cm Conductivity mS cm UO TO SUOIHBIODBHOI 8 ew hepedol g e 90 Q SY oO a 0 S005 10 4 mm 3 CHT 2005 10 4 CHT 2010 3 vs2005 CHT
137. His tag NIF 3 IMAC Profinity IDA A IMAC e 1 IDA NI e 2 Ni IDA e 3 Ni IDA B IMAC NTA C IMAC N Ray Stevens BioLogic DuoFlow BioLogic DuoFlow Maximizer 80 ml min PG 2000 PSI Tronix Inc Teflon FEP 3 2mm 1 6mm IMAC Profinity IMAC
138. 48 Technical Reports SiRNA 50 Miscellanea inside Bio Rad 52 Report 54 54 54 Events Catalog 55 Feedback 56 O Q O O lt HHHMOCUS ConceDpt Gene Expression High Efficiency Low Toxicity High Activity TransFectin Lipid Reagent 120
139. 50 mM KH2PO4 0 3 M KCI 10 6 M pH 8 0 5mM 1 ml Bio Scale 0 2 ml min CV 6 CV 7 10 mM CV 6 550 mM 1 ml B His tag SDS PAGE 1 Precision Plus Protein 2 3 5 IMAC Profinity A 3 B C His tag 6 UJ O O lt O O 29 product GOGOS Flamingo Tom Berkelman
140. 2 o o o 17 0 839 EE 4 HE rm 9 IOA SUOnBIDGHO J 75
141. 2 5 SDS PAGE eFlamingo 532 nm 0 25ng eFlamingo 2 D RUBY eFlamingo MALDI RUBY Flamingo P 16 6 UJ O O O 31 sgus Fr GEARING UP FOR PROTEIN PRODUCTION
142. siRNA B siRNA B siRNA B in vivo siRNA B 3 RNA PCR SM siRNA B 2 2 Pi RNA siRNA
143. 9 ELISA Benchmark Macintosh MacOS
144. RR F Fs re e NW i 1 B RUBY 100001 1265 616 E coli MS 55 25 1394 726 1742 894 g 8g42 5096 1083l519 MA 2265 200 1000 3000 Flamingo 0 25 ng 1g 4 Flamingo 2 E coli RUBY 5 Flamingo 2 D MALDI RUBY 6 5 Flamingo RUBY 2 D E coli 10 gg 2 D 1 ReadyS
145. ir Er mn Mag FI of hit Ce be lm tag 170 9630 1 170 9620 1 1 800 000 980 000 Mag lt dh tenor PT TL MT i IOA SUONeEIPeAO epedolg whatS PCR iQ Multiplex Powermix FAM a Tubulin iQ Multiplex Powermix 1 PCR 1 000 4 PCR cDNA 6 DNA 4 10001 4 10 02 6 1014 18 22 26 30 34 38 42 46 50 HEX GAPDH 100 LIM PlIIIIIIIIIIIIIIII 02 6 1014 1822 26 30 34 38 42 46 50 Texas Red L 1B 1 000 PCR base line subtracted RFU 100
146. http Www bioservers org bioserver Alu HP ALLELE SERVER Manage Groups 1999 Alu 1 1 055 o4 oe 920 oo ooe PV92 PCR Informatics
147. MJ Mini PCR MJ Mini PCR m MiniOpticon PCR MJ Mini 18X32X20cm 4 1kg m LED GD OT NN Web http discover bio rad co jp Practice of the patented polymerase chain reaction PCR process requires a license The MJ Mini thermal cycler and the MiniOpticon system include an Authorized Thermal Cycler and may be used with PCR licenses available from Applied Biosystems Their use with Authorized Reagents also provides a limited PCR license in accordance with the label rights accompanying such reagents Some applications may also require licenses from other third parties Visit us on the Web at discover bio rad co jp
148. Our Focus p 2 12 p22 31 p32 42 p44 48 Technical Reports p50 51 Product Focus Cover Story Bridge a FAX Number 03 5811 6272 genetransfer siRNA siLentFect Lipid Reagent siRNA siLentFect Lipid Reagent siRNA J siHNA 8 1 OD Cycle siRNA siLentFect GAPDH 6well HeLa 10nM GAPDH siRNA siRNA
149. 24 1 b C C 5 mg ml 4 1 a 0 3 ml 100C 5 1 c 5 2 M 0 3 ml 40 1 d CHT Type I 40 m Bio Rad Laboratories 4 0 ID X100mm 40 m 1 mM Sodium phosphate buffer pH6 8 400 mM Sodium phosphate buffer pH6 8 1mM Sodium phosphate 2M guanidi
150. Emission protein 2 e 8 3 EE MS Flamingo Flamingo 512nm 271nm 535nm 0 25 0 5 ng 10 200 ml gt 2 3 40 MS 1XFlamingo 100ml gt 3 2 0 1 w v Tween20 200 ml 10 5 Excitation 512 nm 9X12 cm
151. 2 5 siLentFect 48 CDNA PCR 1009 siLentFect Visit us On the Web at discover Dbio rad cO jD 116 0014 5 18 TEL 03 5811 6270 FAX 03 58 1 1 627 74 Helping Science Serve Mankind Experion rience InnOV Experion RNA View Experion 7 ml Experience
152. 2 5 2 000 ng 1 10 4 1 Pro260 Tris HC1 Experion 1A 2 5 ng 1 10 ng Experion SN 20 1C 1 D 1B Experion Pro260 SDS PAGE Experion
153. 9 IOA SUOnBIDHO 1 1 fee vanced scientific QISC 1 p14 Pharos FX p15 Flamingo p16 PDQuest p17 iQ Multiplex Powermix p18 COS HEKFectin p19 Bio Plex 200 p20 2 BioRadiations A 3
154. BSGCC X Kim 5 Mycoplasma pneumoniae BSGC 10 Kim 5 4
155. IMAC 1 B IMAC 1 mm Pl BLU crmrhr 5 Gryhr 3 300O chr 150 Crrhr 1 0 1 5 BU TD BU GU Run valurm ml 5 IMAC Profinity 75 kD His tag E coli 50 mM 0 3 M NaCl pH 8 0 1 ml 500 mM 280 nm 5 psi 1 IMAC Profinity A 1 60 zm 90zm B IMAC
156. 6 oo or NO ng 3 2 Flamingo SDS PAGE 0 125 980 ng Criterion 4 209 Tris HCI Flamingo A Molecular Imager FX Excitation 532 nm Emission 555 nm longpass B VersaDoc 4000 300nm UV 5 300 nm UV excitation product GO 2500 2000 RS 0 9943 kk OO oO CE9 SD 500 0 1 2 3 4 ng B galactosidase Fluorescence RU 300000 250000 200000 150000 100000 50000 RS 0 9960 0 200 400 600 800 1000 ng B galactosidase A Flamingo ee a w a A Flamingo 1265 613 E coli MS 55 33 870 4226 1299 622 1897728 Intensity 2265 197 eFlamingo
157. PCR PCR RNA PCR 2 MiniOpticon siLentFect Lipid Reagent siRNA PCR ODC ODC RT PCR _
158. Carbonic anhydrase Fluorescence RU 0 a I 0 20 40 60 60 100 ug ml 2 1 50 mM pH 4 0 BSA CA 165 nM Flamingo 510nm 540 nm 2 1 Flamingo 40 200 ml 2 10 Flamingo 100 ml 3 0 1 w v Tween 20 _ 200ml 10 13X9cm 3 8 Flaming9o
159. Copyright reverts to individual authors upon publication Reproraphic copying for personal use is allowed provided credit is given to Bio Rad Laboratories Our Focus Gene Expression 7 Protein Separation 4 Signal Transduction 6 Society and Education 8 See Beyond 10 What s New 14 PharosFX PharosFX Plus 15 Flamingno 16 PDQuest 8 0 Advanced amp Basic 17 iQ Multiplex Powermix 18 COSFectin HEKFectin Cell Specific Lipid 19 Bio Plex PO00 20 Product Focus Experion 22 SDS PAGE IMAC Profinity His tag O Flamingo 3O Cover Story Gearing up for PROTEIN PRODUCTION 00 38 Bridge Explorer 44 SSH 46
160. e ood 9 g J gt Prof Chen Prof Chen Prof Chen Chromatography llove it 1970 30
161. lt 9 EE 1 1 IFA EVOLIS EIA EVOLIS EVOLIS EIA HbA1 Cc Sysmex gt Ai II Ai NG EE i 5 ll 230
162. 2 400 2 800 Flow rate CVhr 2 IMAC 20 mM 50 vv 43 psi 3 bar 20 cm 1 1X30cm Amicon 200 cm h 2 IMAC Profinity 109 psi 7 5 bar 0O UU W gt A QO 9 O product GOGOS 350 5 _ 20 mlM sodium 19 5 phosphate 1 MNaOH 1iMhNa 6Maguardire HH 15 BMM urea 209 EICH 709 EICH FSSEUT S Bed height cm 1B 5 0 gU 400 BOO BOO 1 000 1 2900 FIQw rate Crt 3 IMAC Profinity 20 cm IMAC Profinity 1 1X30cm Amicon His tag
163. 30 19 00 11 O 1B 0 45 0 02 3 74 25 29 160 70 26 00 16 0Q9 16 0 39 0 03 7 0396 a 29 11010 4 900 800 I6 038 003 6 63 50 2g 11990 12 00 10 00 16 0n 38 0 03 5 7496 75 29 127 10 11 00 9 0096 16 0 45 0 03 7 2996 100 2g 105 00 5 00 5 0096 16 0 41 0 03 8 4596 150 2g B7 0O 2 00 3 0096 16 0 26 0 01 5 3496 lculated as 0 CV Experion A Experion B SDS PAGE Pro260 SDS PAGE Pro260 Le marker 4 20 Tris HC1 Experion 100 ng 1 CA 100 ng 1 BSA 3 Experion SDS PAGE 2 Pro260 CV 3 16 SDS PAGE 3 7 31 4
164. throughput fluidics HTF re PC MCV Reagent COOH OO OO rderind Inforrmation Bio Plex 200 Bio Plex Manager 171 000201 Bio Plex 200 171 000201 HIF 171 000205 eo 96Teje Bio Plex 1 Assays luman Mouse Rat TARGETS a A AVAILABLE IL 18 IL 2 I e e ELISA IL 3 EE L 7 IL 8 IL 9 IL 10 IL 12 p40 IL
165. 0 200 TransFectin RNA RT PCR
166. 1 siRNA siRNA siRNA RT PCR RT PCR mRNA mRNA mRNA siRNA siRNA RNA 1 A 1 30 50 confluent
167. 12 p70 IL 13 I L 17 Eotaxin G CSF GM CSF IFN y KC MCP 1 MCAF TM MIP 1e MIP 18 RANTES TNF eg World of Informgtion From a Single Drop Visit us on the Web at discover bio rad cO jDp 116 0014 5 18 TEL 03 5811 6570 FAX 03 58 1 1 627 Helping Science Serve Mankind productOGOS Experion SDS PAGE Karen Zhu Marie Nguyen Wiliam Strong and Christina Whitman Guliaev Bio Rad Laboratories Inc 6000 James Watson Drive Hercules CA 94547 USA SDS PAGE SDS PAGE
168. 43 psi 3 bar CV 3 20 cm 3 43 psi 1 200 cm 1 43 psi
169. 975 Martin The chromatographer should beware of becoming so in love with his techniqune chromatography science art artisSt Chromatography I love it 4 Synergen Synergen Amgen Amgen Colorado 5
170. Applications Tips and TechniqueSs New Products BIOR2OI2IOTS A Resource for Life Science Research Number 006 In this issue Our Focus Experion SDS PAGE IMAC Profinity His tag ETulite Ou 3 Flamingo CBB m 0 25 0 5ng m 2 m 3 m CCD UV MS mm Web http www bio rad con ad flamingo Vis
171. NA siRNA PKR 3 siRNA A B siRNA C siRNA PKR PKR PKR 6 siRNA PKR siRNA PKR siRNA
172. Web mRNA GenBank Accession No Web GenBank BLAST mRNA siRNA Web WP one 0 siRNA B GenBank polymorphism mRNA RNA siRNA in vivo GenBank
173. any Concentration of NaCl in the absence of PO4 Elute with PO4 alone or Combined with NaCI 2 P site C site 2 3 0 M C site DNA IgG 910 suonepeygo g Ta pl TT 20mM NaPO4 pH 6 5 40CV linear gradient to 1M NaCl 20mM PO4 StriD with 0 5M NaPO4 agg
174. d 1 0 ml 4 C 170 3371 49 000 COSFectn CelLSpecific Lipid 1 0 mlX5 4 C 170 3372 158 000 HEKFectin Cell Specific Lipid 0 5 ml 4C 170 3380 32 000 HEKFectin CelLSpecific Lipid 1 0 ml 170 3381 49 000 HEKFectin CelLSpecifc Lipid 1 0 mlX5 4C 170 3382 158 000 whatS NEW Bio Plex 200 Bio Plex Bio Plex 200 Bio Plex Bio PlexTV 200 Bio Plex Manager 4 1 USA FDA 21 CFR Part 11 Bio Plex Manager 4 1 Security Edition SNP Manager Bio Plex High
175. inical Practice cGCP ProteoCell Biotechnologies Vice President of Operations and Scientific Affairs ProteoCell Biotechnologies Inc Laval Quebec Canada Miguel Retamal MSc Retamal 2 000 1 kg 1 10
176. it us On the Web at discover Dbio rad cO jD 116 0014 5 18 TEL 03 581 1 6270 FAX 03 58 1 1 62 Helping Science Serve Mankind 116 0014 5 7 18 TEL 03 5811 6270 532 0025 1 14 11 TEL 06 6308 6568 812 0013 2 18 30 TEL 092 475 4856 OK WE AUND Va WW IE EEAU TEL 03 5811 6271 FAX 03 5811 6272 TEL 03 5811 6626 06 6300 5901 FAX 03 5811 7475 06 6308 3064 On the Cover Conceptual ilustration by Chris Crutchfield ss i ta mi BioHaQdiatOTS PROTEIN PRODUCTION BioRadiations magagine is published by Bio Rad Laboratories Inc 2000 Alfred Nobel Drive Hercules CA 94547 USA 2005 Bio Rad laboratories Inc
177. ne buffer pH6 8 400mM Sodium phosphate 2M guanidine buffer pH6 8 C CHT 2 C CHT C 3 3 C CHT 4
178. regate dna protein A purified human IgG1 3 d A CHT 3 20 mM 1 M NaC1 Aggregates DNA Endotoxin A Purity 1 e Aggregate removal gt 99 by SEC from gt 40 to lt 1 e Leached protein A removal 90 by Cygnus from gt 55 to lt 5ng ml e DNA removal gt 3 logs by PCR down to lt 1ng ml by picogreen e Endotoxin removal 7X10 by LAL down to 1EU ml Cower 1M NaCI 0 5M PO4 25mM PO4 4 3 10 15 25 mM
179. trip IPG 11 cm pH 3 10 NL 2 Criterion 8 16 Tris HCI Flamingo RUBY Molecular Imager FX Excitaton 532 nm Emission 555 nm longpass 6 MALDI 24ng E coli Flamingo RUBY MALDI Proteome Research Services Ann Arbor MI eFlamingo 300 nm UV 0 5ng eFlamingo 3 0 25 960ng eFlamingo
Download Pdf Manuals
Related Search