Screening of Binding Properties of Con-A Immobilized on Bead Cellulose by Flow Microcalorimetry Using Invertase and Anti-Con-A Antibody as Reporting Systems
作者:
P. Dočolomanský,
A. Breier,
P. Gemeiner,
A. Ziegelhöffer,
期刊:
Analytical Letters
(Taylor Available online 1995)
卷期:
Volume 28,
issue 15
页码: 2585-2594
ISSN:0003-2719
年代: 1995
DOI:10.1080/00032719508007411
出版商: Taylor & Francis Group
关键词: Flow micro-calorimeter (Enzyme thermistor);Concanavalin-A;Invertase;Anti-Con-A antibody;Sorbent evaluation
数据来源: Taylor
摘要:
Flow micro-calorimeter so-called enzyme thermistor (ET) is nowadays increasingly used for rapid screening of the sorbents with immobilized protein ligands. In the present paper, chlorotriazine-, dialdehyde-, formylmethyl-, oxirane- and diazotized derivatives of bead cellulose were applied as matrices for immobilization of Concanavalin-A (Con-A). The amounts of free-accessible Con-A in each of the above sorbents were evaluated: i.) by adsorption of invertase; ii.) by means of ELISA using both, mouse anti-Con-A antibody and pig anti-mouse antibody linked with peroxidase. Reactions sub i and ii were monitored in dual way; thermometrically, with the aid of an ET as well as using common spectophotometric methods. The results obtained by spectrophotometry and thermometry intercorrelated significantly (p=0.0028 and p=0.0001, for ELISA and the invertase, respectively). The amounts of immobilized Con-A were directly proportional with the ELISA signal of peroxidase reaction whether this was detected by means of ET (r=0.975, p=0.001) or by spectrophotometry (r=0.983, p=0.0005). Slightly less tight correlation was found between the amounts of immobilized Con-A and the activity of invertase in conjugates when they were assayed by means of thermometry (r=0.933, p=0.0065) and spectrophotometry (r=0.920, p=0.0094). The above data indicate, that thermometry may be considered as a method suitable for rapid evaluation of the amount and accessibility of immobilized Con-A.
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