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The phosphomannan core nanoparticle prepared and characterized in my earlier studies were

lyophilized in order to couple them to the DVS activated seralose matrix and were coupled to

the same, with a long term objective to use this matrix for the purification of mannose 6-

phosphate receptor proteins.

Isolation of Cathepsin D from Unio homogenate and Beta D Glucuronidase from goat

liver tissue.

Unio tissue was homogenized and passed through Lactose affinity chromatography column and

the lactose specific lectin was isolated, the unbound fraction of the homogenate was processed

further and was passed through Pepstatin A affigel column. The bound cathepsin D protein was

eluted from the column using elution buffer. The eluted fractions were concentrated by amicon

concentrator. The sample will be used for nanoparticle preparation and characterized.

Goat liver tissue was homogenized and was processed further to isolate another lysosomal

enzyme Beta D Glucuronidase

Initially the homogenate was passed through Phenyl sepharose column and eluted fractions

were assayed for enzyme activity. Active fractions were pooled and passed through DE-52

(anion exchanger). Unbound sample was removed by extensive washing with column buffer.

Gradient elution was performed for enzyme purification, initially 50mM NaCl in column buffer

was used for elution and then 100 mM and finally 200 Mm

I could find the enzyme activity in 50mM fractions, active fractions were pooled and

concentrated.

To further purify the protein G-200 sephadex matrix was packed in 100ml column and the

partially purified sample was passed through the matrix. The eluate after the void volume were

carefully collected and assayed for enzyme activity and analyzed by SDS-PAGE as shown

below, the band pattern of the enzyme has to be further confirmed

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Preparation and characterization of galactan and galactomannan polysaccharide (from

Strachnous seeds) nanoparticles with Mohammad

Programme of work for next semester

Preparation of nanoparticles for these lysosomal enzymes one prepared from the

invertebrates and one from the vertebrates, and carrying out similar studies as

described in my earlier reports, would further strengthen the role of the receptor in

nanoparticle delivery in specific cell types.

Preparation of antisera for PMC and PMP-BSA in rabbits to quantify the

nanoparticles developed in the study.

In vitro blotting techniques for recognition of nanoparticles by receptor/antibody to

receptor blots Isolation plant polysaccharides (from Moringa oleifera seeds) and preparation of

nanoparticles and characterization of the same.