Tailor-made approach for selective isolation and elution of low-density lipoproteins by immunoaffinity sorbent on silica.
Immunoaffinity procedure was developed for isolation of low density lipoprotein (LDL) from biological samples by using silica-derived immunoaffinity sorbent. Sorbent was prepared by immobilization of monoclonal anti-apoB-100 antibody onto macroporous silica particles, using carefully optimized binding chemistry. Binding capacity of the sorbent towards LDL was determined by batch extraction experiments with solutions of isolated LDL in phosphate-buffered saline, and found to be 8 mg LDL/g. The bound LDL fraction was readily released from the sorbent by elution with ammonia at pH 11.2. The total time needed for isolation procedure was less than 1 h, with LDL recoveries being essentially quantitative for samples containing less than 0.3 mg LDL/mL. With higher concentrations, recoveries were less favorable, most probably due to irreversible adsorption caused by LDL aggreggation. However, reusability studies with isolated LDL at concentration 0.2 mg/mL indicate that the developed immunoaffinity material may be used for multiple binding-release cycles, with minor losses in binding capacity. Finally, the sorbent was successfully applied to isolation of LDL from diluted plasma. Apart from its practical implications for LDL isolation, this study provides crucial insights into issues associated with LDL-sorbent interactions, and may be useful in future efforts directed to development of lipoprotein isolation approaches.
Laboratory of Analytical Chemistry, Department of Chemistry, University of Helsinki, PO. Box 55, FI-00014, Finland.
Ning
N
Gan
Laboratory of Analytical Chemistry, Department of Chemistry, University of Helsinki, PO. Box 55, FI-00014, Finland.
Evgen
E
Multia
Laboratory of Analytical Chemistry, Department of Chemistry, University of Helsinki, PO. Box 55, FI-00014, Finland.
Heli
H
Sirén
Wihuri Research Institute, Haartmaninkatu 8, FI-00290 Helsinki, Finland.
Maija
M
Ruuth
Wihuri Research Institute, Haartmaninkatu 8, FI-00290 Helsinki, Finland.
Katariina
K
Öörni
Laboratory of Analytical Chemistry, Department of Chemistry, University of Helsinki, PO. Box 55, FI-00014, Finland.
Norbert M
NM
Maier
National Institute for Health and Welfare, Genomics and Biomarkers Unit, Biomedicum, FI-00290, Helsinki, Finland.
Matti
M
Jauhiainen
Laboratory of Inorganic Chemistry, Department of Chemistry, University of Helsinki, PO. Box 55, FI-00014, Finland.
Marianna
M
Kemell
Laboratory of Analytical Chemistry, Department of Chemistry, University of Helsinki, PO. Box 55, FI-00014, Finland. Electronic address: marja-liisa.riekkola@helsinki.fi.
Marja-Liisa
ML
Riekkola
D053299:Apolipoprotein B-100 / Q000276:immunology; D002138:Calibration; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007163:Immunosorbent Techniques / Q000295:instrumentation; D008077:Lipoproteins, LDL / Q000302:isolation & purification* / Q000378:metabolism; D017655:Silicon Compounds / Q000737:chemistry; D012822:Silicon Dioxide
D053299:Apolipoprotein B-100; D008077:Lipoproteins, LDL; D017655:Silicon Compounds; D012822:Silicon Dioxide
Low density lipoprotein; Monoclonal anti-apoB-100 antibody; Plasma; Silica based immunosorbent