Biologically produced bifunctional recombinant protein nanoparticles for immunoassays.
Nanoparticles are increasingly used as labels for analytical purposes. In general, nanoparticles need to be functionalized with binding molecules (mostly antibodies or fragments thereof) and label substances using a multistep process that requires several manufacturing and purification steps. Here, we present a biological method of producing functionalized nanoparticles for effective use as label agents in a bioaffinity assay. The particles are based on the globular protein shell of human ferritin. A single chain Fv fragment (scFv) of an antibody is used as the binding moiety and Eu3+ ions as the label substance. Conventional chemical conjugation of the particle and antibody fragment is replaced with genetic fusion between the ferritin subunit and scFv genes. The material, for example, the fusion construct is produced in a single bacterial culture as insoluble forms that are easily purified by centrifugations. The subunits are solubilized and self-assembled, and label ions are introduced by shifting the pH. The functionality of these particles is demonstrated with a bioaffinity assay. This method of producing nanoparticles with inherent antigen binding activity presents several possibilities for the simple production of specific, functional nanoparticles. Production is fast, economical, and environmentally sustainable, making the system advantageous, particularly in applications requiring large quantities of specific nanoparticles.
Jääskeläinen
Anu
A
Department of Biochemistry and Food Chemistry, University of Turku, 20014 Turku, Finland.
Harinen
Reija-Riitta
RR
Soukka
Tero
T
Lamminmäki
Urpo
U
Korpimäki
Teemu
T
Virta
Marko
M
D000911:Antibodies, Monoclonal / Q000276:immunology; D001665:Binding Sites; D002412:Cations / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D005063:Europium / Q000737:chemistry; D005293:Ferritins / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007118:Immunoassay / Q000379:methods; D007128:Immunoglobulin Fragments / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D053758:Nanoparticles / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism*; D013972:Thyrotropin / Q000737:chemistry / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D002412:Cations; D007128:Immunoglobulin Fragments; D011994:Recombinant Proteins; C055966:immunoglobulin Fv; D005063:Europium; D013972:Thyrotropin; D005293:Ferritins