Disc Electrophoresis – Ii Method and Application to Human Serum Proteins*

Davis, B.

doi:10.1111/j.1749-6632.1964.tb14213.x

Cited by 17,879 publications

(822 citation statements)

References 9 publications

Supporting

Mentioning

743

Contrasting

Unclassified

Order By: Relevance

“…For electrophoresis, 6 ll of serum samples was incubated with 1 ll of 10% haemolysate of washed human red blood cells to enable the Hb content of the haemolysate to form Hp-Hb complex, after which 6 ll of loading buffer was added to the reaction mixture. Then 8 ll of each reaction mixture was loaded for electrophoresis according to the method described by Davis [36] after which proteins were fixed using 10% trichloroacetic acid solution and stained with benzidine solution. The bands were then observed for Hp phenotyping.…”

Section: Haptoglobin Phenotypingmentioning

confidence: 99%

Haptoglobin Polymorphism and Association with Complications in Ghanaian Type 2 Diabetic Patients

et al. 2011

View full text Add to dashboard Cite

There is scanty information on the role of genetic factors, especially those relating to haptoglobin (Hp) phenotypes in the expression of complications among diabetes mellitus patients in Ghana. In this study, we investigated whether there is any association between Hp phenotypes and diabetic complications and to determine if association of the Hp phenotypes with diabetic complications in Ghanaian diabetics differ from those in Caucasians. A total of 398 participants were randomly recruited into the study. These comprised diabetic patients numbering 290 attending a diabetes Clinic in Ghana and 108 non-diabetic controls from the same community. Analyses of the results indicate that most of the diabetics with complications were of the Hp 2-2 (35%) and Hp 2-1 (23.9%) phenotypes. Fewer diabetics were found to be of the Hp 2-1 M phenotype. The controls were mostly of Hp 1-1 and Hp 2-1 M phenotypes. The odds ratio of having complications in a diabetic with an Hp 2-2 phenotype was 18.27 times greater than that for Hp 0-0. Hp 2-2 phenotype with its poor antioxidant activity may therefore be a useful predictor for the propensity of an individual to develop diabetes complications.

show abstract

Section: Haptoglobin Phenotypingmentioning

confidence: 99%

Haptoglobin Polymorphism and Association with Complications in Ghanaian Type 2 Diabetic Patients

et al. 2011

View full text Add to dashboard Cite

show abstract

“…Polyacrylamide gel electrophoresis (PAGE) was performed as described by Davis [40], and SDS-PAGE according to Laemmli [41]. For native PAGE, the gel was run at 100 V for 2.5 h, and the molecular mass markers used were thyroglobulin (669 kDa), ferritin (440 kDa), catalase (232 kDa), lactate dehydrogenase (140 kDa) and bovine serum albumin (66 kDa) (Pharmacia).…”

Section: Electrophoresis and Enzyme Stainingmentioning

confidence: 99%

Purification and characterisation of phenoloxidase from amphioxus Branchiostoma belcheri tsingtauense

Pang

Zhang

Shi

et al. 2005

Fish & Shellfish Immunology

View full text Add to dashboard Cite

“…A small amount may also be contributed by cystines; i.e., cysteine sidechains in a disulfide linkage (each cysteine in such a disulfide structure is a half cystine). The standard method of denaturing proteins and separation by electrophoresis in acrylamide is used [19][20][21][22]. Although the LuxG expressed in P. leiognathi has been shown to function as a homodimer, it migrates as a monomer on a denaturing gel.…”

Section: Exercise 4: Luxg-his 6 Concentration By Direct Spectrophotommentioning

confidence: 99%

Expression, purification, and characterization of a recombinant flavin reductase from the luminescent marine bacterium Photobacterium leiognathi

Crowley

2010

Biochem Molecular Bio Educ

View full text Add to dashboard Cite

In Photobacterium, the flavin reductase encoded by luxG regenerates the reduced form of flavin mononucleotide (FMN). Reduced FMN is one of the substrates of the luciferase enzyme that catalyzes a lightemitting reaction. A set of experiments, that employs a luxG-expression plasmid construct (pGhis) and is suitable for an undergraduate laboratory course, is presented. Hexahistidine-tagged protein is expressed in E. coli from pGhis, with the T7 RNA polymerase/lac repressor induction system. Bacteria are lysed by sonication and the tag allows for purification by immobilized metal ion affinity chromatography. A gel filtration column is used to remove ions and the other small molecules. The Bradford assay, with multiwell plates and an automated plate reader, is used to identify protein concentration peaks from both columns. The concentration of purified enzyme is then calculated from its A 280 using the predicted extinction coefficient. Yield and purity are further assayed with SDS-PAGE. Activity of purified enzyme is measured with riboflavin or FMN as substrate. Reaction rate is quantified by monitoring decrease in A 340 as the redox partner, NADH, is oxidized.Keywords: recombinant protein expression, immobilized metal ion affinity chromatography, LuxG, flavin reductase.Bioluminescence occurs in a variety of organisms; e.g., firefly, marine invertebrates and bacteria. Most of the luminescent bacteria exist in symbiosis with fish, proliferating within certain tissues and making the fish appear to be luminescent. The reaction that generates the light energy is catalyzed by the luciferase enzyme in all these organisms. However, the substrates of the reaction vary quite a bit. In the luminescent marine bacteria Photobacterium leiognathi and Photobacterium fischeri (previously known as Vibrio fischeri), one of the substrates is the fully-reduced, quinone form of flavin mononucleotide (FMN):As the light-generating reaction involves oxidation of FMNH 2 to FMN, sustained luminescence necessitates the recycling of FMN back to the reduced form. Such a reaction (Fig. 1) is usually catalyzed by a flavin reductase enzyme [2] (the reduced form is often shown as FMNH 2 , however at pH > 6.3 it ionizes to FMNH 2 [3]). NADH is oxidized as FMN is reduced.In P. fischeri and P. leiognathi the luciferase enzyme is an ab heterodimer [4]. The genes for these subunits, luxA and luxB, are encoded within the lux operon [5][6][7][8]. This operon also includes luxG that codes for a flavin reductase (protein is designated LuxG [9]). Thus, the translation of the lux operon-encoded mRNA should generate equimolar amounts of luciferase enzyme and flavin reductase.

show abstract

Disc Electrophoresis – Ii Method and Application to Human Serum Proteins*

Abstract: Summary The technique of disc electrophoresis has been presented, including a discussion of the technical variables with special reference to the separation of protein fractions of normal human serum.

Cited by 17,879 publications

References 9 publications

Haptoglobin Polymorphism and Association with Complications in Ghanaian Type 2 Diabetic Patients

Haptoglobin Polymorphism and Association with Complications in Ghanaian Type 2 Diabetic Patients

Purification and characterisation of phenoloxidase from amphioxus Branchiostoma belcheri tsingtauense

Expression, purification, and characterization of a recombinant flavin reductase from the luminescent marine bacterium Photobacterium leiognathi

Contact Info

Product

Resources

About