ChromoBlog

Introduction

Blocking is an essential step during the preparation of a sample for immunofluorescence detection. Blocking improves the sensitivity by reducing nonspecific background and therefore increases image quality. Insufficient blocking results in higher background noise and over-blocking can even mask the specific signal.

Simultaneous immunostaining, also called one-step immunostaining vs. sequential immunostaining. Nano-Secondaries stain different primary antibodies equally well in one-step staining and sequential staining.

HeLa cells were immunostained with different primary antibodies and Nano-Secondaries Alexa Fluor® 647 (1:1,000, magenta). Cell nuclei were stained with DAPI (blue). Scale bar, 20 μm.

Nano-Secondaries are monoclonal Nanobodies that bind to primary antibodies in a species and subtype specific manner, i.e. Nano-Secondaries are very precise secondary antibodies. They are recombinantly produced in bacterial or other animal-free expression systems; in contrast, classical mono- or polyclonal secondary antibodies are traditionally produced using hybridoma technologies or are isolated from a host’s blood.