Stabilization of proteins immobilized on Sepharose from leakage by glutaraldehyde crosslinking

Kowal, Robert C.; Parsons, Robert G.

doi:10.1016/0003-2697(80)90319-x

Cited by 78 publications

(15 citation statements)

References 9 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…28 We examined the effect of glutaraldehyde, a dialdehyde, on immobilized HSA. A reagent of this type should be capable of linking two adjacent HSA molecules together via their primary amine groups.…”

Section: Discussionmentioning

confidence: 99%

The influence of bond chemistry on immobilized enzyme systems for ex vivo use

Comfort¹,

Mullon²,

Langer³

1988

Biotech & Bioengineering

View full text Add to dashboard Cite

The ideal derivatized support for the clinical use of an immobilized enzyme system should irreversibly bind active enzyme. We have investigated the behavior of heparinase and bilirubin oxidase immobilized via cyanogen bromide, tresyl chloride, epoxide, or carbodiimidazole activated natural and synthetic matrices. The protein bound to each activated support was 90% for cyanogen bromide (CNBr) activated agarose, 50-80% for tresyl chloride activated agarose, and 50% for oxirane activated acrylic (Eupergit C). The activity retention of immobilized heparinase was greatest (50%) with CNBr activated agarose while for the immobilization of bilirubin oxidase, the activity retention was greatest (25-30%) with tresyl chloride activated agarose and oxirane activated acrylic.The stability of the different covalent bonds was studied in vitro with radioiodinated enzymes. The leaching profiles showed the same trends for each support and chemistry. A plateau in protein leaching was reached after a few hours of incubation and the transient leaching period was well represented by a logarithmic function of time. The amount of enzyme released from the least stable support (CNBr activated agarose) in 24 h was injected intravenously in New Zealand white rabbits. Using an indirect enzyme-linked immunosorbant assay (ELISA), no immune response was detected. The transient leaching profile was shortened by washing the enzyme-support conjugate with 1M hydroxylamine, pH8.5 intermolecular cross-linking with glutaraldehyde also improves the enzyme-support stability. Tresyl chloride and oxirane activated supports produce bonds with improved stability without adversely affecting enzymatic activity.

show abstract

Section: Discussionmentioning

confidence: 99%

The influence of bond chemistry on immobilized enzyme systems for ex vivo use

Comfort¹,

Mullon²,

Langer³

1988

Biotech & Bioengineering

View full text Add to dashboard Cite

show abstract

“…The yeast Saccharomyces cerevisiae was obtained from a commercial store (Angel Yeast Co., Ltd., China). The affinity sorbent for crosslinking albumin with PSM was prepared according to the method described previously (Kowal and Parsons, 1980;Belogortseva et al, 1994).…”

Section: Methodsmentioning

confidence: 99%

Purification of a secreted lectin from Andrias davidianus skin and its antibacterial activity

Tong

Kong

et al. 2015

Comparative Biochemistry and Physiology Part C: Toxicology &

View full text Add to dashboard Cite

“…Expression and purification of GST-fusion protein using glutathione-Sepharose 4B beads (Pharmacia) was performed using described methods [20]. An affinity column with golgin-67 protein was prepared by glutaraldehyde coupling method [21] and used to affinity purify rabbit antibodies as previously described in standard methods [22]. Purified antibodies were analysed by indirect immunofluorescence.…”

Section: Affinity Purification Of Antibodymentioning

confidence: 99%