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1. (WO2013163451) STAIN-FREE PROTEIN QUANTIFICATION AND NORMALIZATION
Latest bibliographic data on file with the International Bureau   

Pub. No.: WO/2013/163451 International Application No.: PCT/US2013/038250
Publication Date: 31.10.2013 International Filing Date: 25.04.2013
IPC:
G01N 21/64 (2006.01) ,G01N 21/76 (2006.01) ,G01N 21/33 (2006.01) ,G01N 33/52 (2006.01) ,G01N 33/68 (2006.01) ,G06F 19/20 (2011.01) ,G01J 1/18 (2006.01) ,G01J 3/443 (2006.01) ,C12Q 1/06 (2006.01)
G PHYSICS
01
MEASURING; TESTING
N
INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
21
Investigating or analysing materials by the use of optical means, i.e. using infra-red, visible, or ultra-violet light
62
Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
63
optically excited
64
Fluorescence; Phosphorescence
G PHYSICS
01
MEASURING; TESTING
N
INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
21
Investigating or analysing materials by the use of optical means, i.e. using infra-red, visible, or ultra-violet light
75
Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
76
Chemiluminescence; Bioluminescence
G PHYSICS
01
MEASURING; TESTING
N
INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
21
Investigating or analysing materials by the use of optical means, i.e. using infra-red, visible, or ultra-violet light
17
Systems in which incident light is modified in accordance with the properties of the material investigated
25
Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
31
Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry
33
using ultra-violet light
G PHYSICS
01
MEASURING; TESTING
N
INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
33
Investigating or analysing materials by specific methods not covered by groups G01N1/-G01N31/131
48
Biological material, e.g. blood, urine; Haemocytometers
50
Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
52
Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper
G PHYSICS
01
MEASURING; TESTING
N
INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
33
Investigating or analysing materials by specific methods not covered by groups G01N1/-G01N31/131
48
Biological material, e.g. blood, urine; Haemocytometers
50
Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
68
involving proteins, peptides or amino acids
G PHYSICS
06
COMPUTING; CALCULATING; COUNTING
F
ELECTRIC DIGITAL DATA PROCESSING
19
Digital computing or data processing equipment or methods, specially adapted for specific applications
10
Bioinformatics, i.e. methods or systems for genetic or protein-related data processing in computational molecular biology
20
for hybridisation or gene expression, e.g. microarrays, sequencing by hybridisation, normalisation, profiling, noise correction models, expression ratio estimation, probe design or probe optimisation
G PHYSICS
01
MEASURING; TESTING
J
MEASUREMENT OF INTENSITY, VELOCITY, SPECTRAL CONTENT, POLARISATION, PHASE OR PULSE CHARACTERISTICS OF INFRA-RED, VISIBLE OR ULTRA-VIOLET LIGHT; COLORIMETRY; RADIATION PYROMETRY
1
Photometry, e.g. photographic exposure meter
10
by comparison with reference light or electric value
16
using electric radiation detectors
18
using comparison with a reference electric value
G PHYSICS
01
MEASURING; TESTING
J
MEASUREMENT OF INTENSITY, VELOCITY, SPECTRAL CONTENT, POLARISATION, PHASE OR PULSE CHARACTERISTICS OF INFRA-RED, VISIBLE OR ULTRA-VIOLET LIGHT; COLORIMETRY; RADIATION PYROMETRY
3
Spectrometry; Spectrophotometry; Monochromators; Measuring colours
28
Investigating the spectrum
443
Emission spectrometry
C CHEMISTRY; METALLURGY
12
BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
Q
MEASURING OR TESTING PROCESSES INVOLVING ENZYMES OR MICRO-ORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
1
Measuring or testing processes involving enzymes or micro-organisms; Compositions therefor; Processes of preparing such compositions
02
involving viable micro-organisms
04
Determining presence or kind of micro-organism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor
06
Quantitative determination
Applicants:
BIO-RAD LABORATORIES, INC. [US/US]; 1000 Alfred Nobel Drive Hercules, CA 94547, US
Inventors:
FREEBY, Steve; US
LIU, Ning; US
MCDONALD, Kevin; US
PAULUS, Aran; US
POSCH, Anton; DE
Agent:
ANTHONY, Peter; Kilpatrick Townsend and Stockton LLP 2 Embarcadero Center, 8th Floor San Francisco, CA 94111, US
Priority Data:
61/639,68627.04.2012US
61/639,69227.04.2012US
Title (EN) STAIN-FREE PROTEIN QUANTIFICATION AND NORMALIZATION
(FR) DÉTERMINATION QUANTITATIVE ET NORMALISATION DE PROTÉINES SANS COLORATION
Abstract:
(EN) Disclosed herein are methods of protein quantification and normalization using haloalkylated tryptophan fluorescence. Complex protein samples, i.e., samples that each contain 1,000 or more distinct proteins, from diverse sources that do not have common protein profiles are treated with a halo-substituted organic compound (i.e. haloalkane) that reacts with tryptophan residues to form fluorescent products. Irradiation of the samples with ultraviolet light and the detection and quantification of the resultant fluorescent emissions from all proteins in each sample are then used to obtain comparative values for total protein content among the various samples. The values thus obtained are found to be valid indications of comparative total protein content, despite the fact that the tryptophan levels vary widely among the various proteins in any single sample and the samples, due to the diversity of their origins, tend to differ among themselves in the identities and relative amounts of the proteins that they contain. Protein samples are also normalized to correct for differences in sample dilution, sample loading, and protein transfer inconsistencies, by using stain-free detection of total protein in each of the samples, or detection of subsamples within each sample.
(FR) La présente invention concerne des procédés de détermination quantitative et de normalisation de protéines à l'aide de la fluorescence du tryptophane halogénoalkylé. Des échantillons de protéines complexes, à savoir des échantillons qui contiennent chacun 1 000 protéines distinctes ou plus, de diverses sources qui ne présentent pas de profils protéiques communs, sont traités avec un composé organique à substitution halo (c'est-à-dire halogénoalcane) qui réagit avec des résidus tryptophanes pour former des produits fluorescents. L'irradiation des échantillons avec un rayon ultraviolet et la détection et la détermination quantitative des émissions fluorescentes obtenues de toutes les protéines dans chaque échantillon sont alors utilisées pour obtenir des valeurs comparatives pour la teneur totale en protéines parmi les divers échantillons. Les valeurs ainsi obtenues s'avèrent être des indications valables de la teneur totale en protéines comparative, malgré le fait que les concentrations de tryptophane varient largement parmi les diverses protéines dans n'importe quel échantillon unique et que les échantillons, en raison de la diversité de leurs origines, ont tendance à différer entre eux dans les identités et les quantités relatives des protéines qu'ils contiennent. Les échantillons de protéines sont également normalisés pour corriger les différences de dilution d'échantillon, de charge d'échantillon et d'incohérences de transfert de protéines, en utilisant la détection sans coloration des protéines totales dans chacun des échantillons, ou la détection de sous-échantillon au sein de chaque échantillon.
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Designated States: AE, AG, AL, AM, AO, AT, AU, AZ, BA, BB, BG, BH, BN, BR, BW, BY, BZ, CA, CH, CL, CN, CO, CR, CU, CZ, DE, DK, DM, DO, DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, HN, HR, HU, ID, IL, IN, IS, JP, KE, KG, KM, KN, KP, KR, KZ, LA, LC, LK, LR, LS, LT, LU, LY, MA, MD, ME, MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ, OM, PA, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SC, SD, SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM, TN, TR, TT, TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, ZW
African Regional Intellectual Property Organization (ARIPO) (BW, GH, GM, KE, LR, LS, MW, MZ, NA, RW, SD, SL, SZ, TZ, UG, ZM, ZW)
Eurasian Patent Office (AM, AZ, BY, KG, KZ, RU, TJ, TM)
European Patent Office (EPO) (AL, AT, BE, BG, CH, CY, CZ, DE, DK, EE, ES, FI, FR, GB, GR, HR, HU, IE, IS, IT, LT, LU, LV, MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, SI, SK, SM, TR)
African Intellectual Property Organization (BF, BJ, CF, CG, CI, CM, GA, GN, GQ, GW, ML, MR, NE, SN, TD, TG)
Publication Language: English (EN)
Filing Language: English (EN)