Purification Of Human Growth Hormone Research Articles

Design of peptide ligands for affinity purification of human growth hormone

AbstractBackgroundIn this study, we have demonstrated the design, screening and selection of peptide ligands for the affinity capture of human growth hormone (hGH) from yeast cell cultures.ResultsLigand design was carried out using multiple approaches based on primary sequence and structures of natural binding partners of hGH. Screening of potential affinity peptides was conducted using high throughput microarray platforms followed by assessment of in‐solution binding to hGH using fluorescence polarization. Peptide leads were subsequently immobilized on chromatographic resins and the binding and desorption behavior was examined using batch adsorption studies. A lead candidate was examined in further details in column chromatography studies which indicated that while high purity was attained, further refinement was necessary for improved product recovery. Histidine scanning was employed to successfully improve the recovery of hGH from cell culture fluid while still maintaining high purity. Finally, proof‐of‐concept was demonstrated in the column format using complex feed stock where a product purity of 95% was attained at 80% yield.ConclusionThe approaches presented here can be translated to other biologics of interest for the rapid development of affinity based purification processes. © 2019 Society of Chemical Industry

Small-Molecule Affinity Ligands for Protein Purification: Combined Computational Enrichment and Automated In-line Screening of an Optically Encoded Library

Screening with codes: A versatile methodology for the development of small-molecule affinity chromatography ligands for protein targets is presented. The combination of computational enrichment with automated screening and affinity mapping of an optically encoded combinatorial library allows identification of a novel set of ligands for the single-step purification of human growth hormone (see picture).

Immunoadsorptive purification of human growth hormone

Immunoadsorptive purification of human growth hormone

Purification of pituitary and biosynthetic human growth hormoneusing monoclonal antibody immunoadsorbent

This report describes the purification of human growth hormone from crude pituitary extract and lysate of recombinant E. coli by an immunoadsorbent purification with monoclonal antibody coupled to solid phase. By a single-immunoaffinity chromatography step pure hGH can be obtained from either origin as revealed by SDS-PAGE followed by silver staining or immunoblotting. An additional ion-exchange chromatography step results in homogeneous 22 kDa hGH preparations. Furthermore, this method may be used for isolation of a pituitary hGH variant which has higher binding affinity for this monoclonal antibody than the major 22 kDa form. This study clearly illustrates the potential of monoclonal antibody immunoadsorbents for purification of different molecular forms of hGH.

Radioiodination of human growth hormone with characterization and minimization of the commonly defined “damaged products”

The radioiodination and Chromatographie purification of human growth hormone (hGH) has been studied in order to better define and control the so-called “preparation damage”, which is often a cause of interferences, loss in specific activity and sensitivity, misclassification errors in radioligand assays, and a source of misinterpretation when the tracer is used in receptors or in vivo studies. A series of labelings and false labelings, with and without protein carrier in the buffer used for Sephadex purification, indicate that the “preparation damage” peak is made up of two components: aggregated 125I-hGH and BSA-carried radioactivity. The former can be minimized by the use of recently extracted non-lyophilized hGH, and the latter by enzymatic labeling. Both components can be better resolved, and thus eliminated, when Sephadex G-100 is employed rather than G-75.

A simple method for the extraction and purification of human growth hormone and its assay by paper chromatography.

ABSTRACT A simple method for the extraction and purification of human growth hormone has been developed, by which, after the initial alkaline extraction of the acetone-dried pituitary powder, the bulk of the impurities is precipitated at pH 4.8 and the crude hormone subsequently precipitated with alcohol at 25% v/v concentration. The crude hormone is then purified by a single Sephadex G-100 gel nitration, 75% of the activity of the crude being obtained as virtually pure monomer and the remaining 25% in the form of two fractions containing two separate polymers. A simple and rapid paper chromatographic assay method has also been developed for following the extraction and purification stages which gives good agreement with the corresponding radioimmunoassays.

Improved method for the extraction and purification of human growth hormone.

A simple method has been devised for the extraction and purification of human growth hormone (HGH) from acetone-dried pituitary glands. The HGH, suitable for clinical use, is recovered in one fraction in a yield of 70–80 IU/g of pituitary powder. A step is included to preserve the glycoprotein hormones in the glands. The method can also be applied to extracting clinically useful HGH from embalmed pituitary glands in a yield of 13–22 IU/g of pituitary powder. For chemical, immunochemical or special clinical uses, HGH may be purified further by chromatography on DEAE-cellulose.

Purification of human growth hormone from pre-extracted pituitary residues.

Purification of human growth hormone from pre-extracted pituitary residues.

Heterogeneity of human growth hormone preparations by immuno-gel filtration and gel filtration electrophoresis.

VARIOUS methods have been used for the purification of human growth hormone (HGH). The method described by Roos et al.1 uses especially mild conditions with extraction of homogenized frozen pituitary glands at pH 6.2, followed by ammonium sulphate precipitation and gel filtration on ‘Sephadex’. We sought to check the homogeneity of preparations obtained by this method which had been lyophilized and dispensed for therapeutic use. With the small amounts of material available it seemed convenient to apply thin-layer gel filtration2 on ‘Sephadex G-200’ in combination with immunodiffusion analysis (immuno-gel filtration) or with electrophoresis (gel filtration electrophoresis)3,4.

Purification of human growth hormone on "sephadex G-200".

WE have previously reported the preparation of human growth hormone (HGH) from acetone powders of pituitary glands1. The hormone preparations were obtained in high yield and proved satisfactory in clinical investigations but were by no means electrophoretically homogeneous. We wish to describe now a simple and reproducible method for further purification of this material which has resulted in a growth hormone preparation of markedly increased specific biological activity. This preparation, although still not homogeneous, was shown to be substantially purified by the criteria of disk electrophoresis and ultracentrifugation. No serum protein contamination could be detected when this purified preparation was assayed by immunological techniques. A second fraction obtained by this procedure showed some growth hormone activity and in addition possessed considerable luteinizing hormone (LH) potency. This fraction was analysed by immunological techniques and found to contain serum albumin and a smaller amount of serum globulins.