Abstract
This monograph summarizes recent developments in the purification and analysis of natural and recombinant proteins. The basic strategies employed in protein purification are reviewed with regards to the characteristics of the protein of interest that may aid its isolation, choice of the starting material, and use of denaturants. Preparation of cell-free extracts followed by bulk precipitation and/or phase partition constitute the initial steps of many purification schemes. Chromatographic methods (size exclusion, ion exchange, hydroxylapatite, reversed phase, hydrophobic interaction and affinity based) utilizing either traditional, low pressure or high-performance liquid chromatography instrumentation are discussed. Electrophoretic techniques used to analyze the homogeneity of the protein product include SDS-PAGE, isotachophoresis, IEF and two dimensional gel electrophoresis.
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Keywords
- Sodium Dodecyl Sulfate
- Size Exclusion Chromatography
- Protein Purification
- Purification Scheme
- Hydrophobic Interaction Chromatography
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.
Abbreviations
- a:
-
selectivity factor
- ABx:
-
antibody exchanger
- cDNA:
-
complementary DNA
- CM:
-
carboxymethyl
- CoA:
-
coenzyme A
- D:
-
diameter
- Da:
-
Daltons
- DEAE:
-
diethylaminoethyl
- DNA:
-
deoxyribonucleic acid
- FPLC:
-
fast protein liquid chromatography
- FR:
-
flow rate
- HETP:
-
height equivalent to a theoretical plate
- HIV-1:
-
human immunodeficiency virus type 1
- HPLC:
-
high-performance liquid chromatography
- IEF:
-
isoelectric focusing
- IgG:
-
immunoglobulin G
- IgM:
-
immunoglobulin M
- K:
-
distribution coefficient
- k′:
-
capacity factor
- L:
-
length
- LV:
-
linear velocity
- N:
-
number of theoretical plates
- PAGE:
-
polyacrylamide gel electrophoresis
- PEI:
-
polyethyleneimine
- PEG:
-
polyethylene glycol
- pI:
-
isoelectric point
- PMSF:
-
phenylmethylsulfonyl fluoride
- QAE:
-
quaternary aminoethyl
- R:
-
resolution
- RNA:
-
ribonucleic acid
- Rt:
-
retention time
- SDS:
-
sodium dodecyl sulfate
- SF:
-
scale-up factor
- SP:
-
sulfoxypropyl
- TFA:
-
trifluoroacetic acid
- V:
-
retention volume
- V0 :
-
void volume
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© 1989 Springer-Verlag
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Lillehoj, E.P., Malik, V.S. (1989). Protein purification. In: Bioprocesses and Engineering. Advances in Biochemical Engineering/Biotechnology, vol 40. Springer, Berlin, Heidelberg. https://doi.org/10.1007/BFb0009827
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DOI: https://doi.org/10.1007/BFb0009827
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