Thin layer isoelectric focusing of the soluble lens extracts from larval stages and adult Xenopus laevis

Brahma, S. K.; Bours, J.

doi:10.1016/0014-4835(72)90112-1

Cited by 22 publications

(6 citation statements)

References 16 publications

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“…They prepared antisera to the total proteins of adult frog lenses and to a gamma crystallin fraction of these lenses obtained by chromatography on D EAE-cellulose and Sephadex G-100. Isoelectric focusing of lens extracts from larval and adult Xenopus laevis also demonstrated a gradual decrease in gamma crystallins and an increase in beta crystallins in lenses from older animals and a large increase in alpha crystallins after metamorphosis (BRAHMA and BOURS, 1972). Antibodies to total lens proteins were absorbed with mouse tissue powder and frog liver powder before use.…”

Section: Lens Induction and Cytodifferentiationmentioning

confidence: 99%

The Amphibian Eye: Development and Regeneration

Reyer

1977

The Visual System in Vertebrates

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Section: Lens Induction and Cytodifferentiationmentioning

confidence: 99%

The Amphibian Eye: Development and Regeneration

Reyer

1977

The Visual System in Vertebrates

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“…Isoelectric focusing experiments were carried out as described in previous work (BOURS, 1971 ;BRAHMA & BOURS, 1972;BOURS, 1973a, c;BOURS & BRAHMA, 1973). Immunoelectrophoresis experiments were carried out as described previously (BOURS, 1973a).…”

Section: Methodsmentioning

confidence: 99%

“…Another technique to determine contaminating proteins is (4) the injection of the purified protein by preparing antibodies in rabbits. The normal subcutaneous injections can be administered, or the lymph node injection method (BOORS & VAN DOORENMAALEN, 1972;BRAHMA et al, 1972) can be used in case of minute amounts of antigens available (< 1 mg). By polyacrylamide thin-layer isoelectric focusing, contaminating proteins will reveal themselves as one or more extra protein bands of other isoelectric points than the protein isolated.…”

Section: Examhlation Of the Purity Of Proteins Isolated By Free Isoelmentioning

confidence: 99%

Isoelectric focusing and immunochemistry of lens crystallins

Bours

1974

Doc Ophthalmol

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This is a review of 11 published articles with some conclusive results on the isoelectric focusing and immunochemistry of lens crystallins. The techniques used throughout the work are isoelectric focusing and immunological methods.The isoelectric focusing technique is performed on an analytical scale to realize the separation of proteins by isoelectric focusing in thin-layer polyacrylamide gels. The importance of carrying out all isofocusing techniques at 4 ~ C, and also of measuring the pH gradients established at the same temperature is noticed. Free isoelectric focusing is used as a preparative step to isolate uncontaminated crystallin fractions. Both analytical and preparative isoelectric focusing experiments of crystallins performed throughout the work are in absence of urea or any other dissociating medium.The purity of proteins isolated by free iseelectric focusing is examined by criteria of homogeneity which are founded on three different techniques, e.g. thin-layer isoelectric focusing, immunoelectrophoresis and antigen/antibody crossed electrophoresis.The existence of various crystallins in the chick lens, like ~z-, ~-crystallin and FISC, related to own data, is reviewed and nomenclatures of chick lens crystallins are compared with present data from literature. The antigenic properties of proteins from chick intra-ocular tissues are also discussed. Anti-total iris serum contains antibodies against crystallins and serum proteins. The presence of proteins immunologically identical to lens crystallins in chick iris is shown by indirect immunofluorescence experiments: brilliant fluorescence of the lens is observed due to the presence of crystallin antibodies in anti-total iris serum. It is concluded that proteins of all intra-ocular tissues which are immunologically identical to lens crystallins are essential structural elements. Contrary to this statement several authors support the theory that the presence of proteins immunologically identical to lens crystallins in the intra-ocular tissues is due to a post-mortem diffusion of these antigens from the lens. The presence of several erum proteins in all intra-ocular tissues excepting lens is shown. Evidence is presented that the number of serum proteins in intra-ocular tissues is restricted by the various blood/ocular barriers, and that the blood/retinal barrier is of the same nature as the blood/brain barrier.The presence ~-, ~-, ,,,-and pre-~-crystallins is shown in various stages of the bovine embryonic lens by thin-layer isoelectric focusing and various immunological methods. It is defined that the bovine -~,-crystallins have isoelectric points higher than pI --7.0. It is shown by immuno-osmophoresis that the embryonic ~-crystallins consist of two molecular forms with different electrophoretic mobilities. The ~-crystallins are forming half the total amount of the crystallins present in the embryonic lens. It is concluded that the "c-crystallins are more prominent in embryonic cow lens and in adult lens nucleus and less prominent in adult lens cortex. The...

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“…Calculation of crystallin moieties of total water-soluble crystallins (in %) after densitometry was done according to Brahma and Bours [1972]. The standard error of the thin-layer IE F proce dure with densitometric evaluation is ± 4.6%.…”

Section: Methodsmentioning

confidence: 99%

Biochemistry of the Ageing Rat Lens

Bours¹,

Hockwin²

1983

Ophthalmic Res

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Thin-layer isoelectric focusing of the water-soluble crystallins shows with increasing age of the animals an increase of α-crystallins of high and low molecular weight and of the total β-crystallins content. The β-crystallin components of high molecular weight also increased during ageing, but the β-crystallins of lower molecular weight decreased considerably. The total γ-crystallin content, as well as certain γ-crystallin components also decreased considerably with increasing age, or vanished completely. From 429 to 1,211 days of age the male γ-crystallins as percent of the water-soluble moiety, were higher than the female γ-crystallins. As a consequence of ascending age, the content of water-insoluble proteins of the rat lens increased gradually and continuously, e.g. by a process of insolubilization of soluble β- and γ-crystallins.

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Thin layer isoelectric focusing of the soluble lens extracts from larval stages and adult Xenopus laevis

Cited by 22 publications

References 16 publications

The Amphibian Eye: Development and Regeneration

The Amphibian Eye: Development and Regeneration

Isoelectric focusing and immunochemistry of lens crystallins

Biochemistry of the Ageing Rat Lens

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