This Imaging Bites seminar will present OneStep-IF - OneStep-IF uses secondary nanobodies (“SmartSecondaries®”) premixed with primary antibodies prior to sample incubation, forming defined Ab::Nb complexes in a single staining step, thereby removing the classical multiplexing bottleneck in immunofluorescence.

This premixing strategy separates antibody specificity from species constraints, enabling simultaneous use of multiple primary antibodies from the same host without cross-reactivity (e.g., multiple mouse monoclonal primaries). Using nanobodies as secondary binders reduces linkage error, improves epitope accessibility, and provides stable, stoichiometric labeling. As a result, OneStep-IF combines simplicity, robustness, and quantitative control, making it compatible with nearly any antibody-based technique.

This concept enables scalable multiplexing across imaging modalities. In NanoPlex (Mougios et al., Nature Communications 2024), OneStep-IF allows the detection of 21 protein targets in a single neuronal sample with high specificity using standard confocal microscopy. SUM-PAINT builds on the same principle to achieve 30-plex, 3D super-resolution imaging at single-protein resolution, by linking SmartSecondaries to DNA barcodes and a fast DNA-PAINT readout, enabling spatial proteomics beyond traditional immunofluorescence (Unterauer et al. Cell 2024)

Together, these examples demonstrate that OneStep-IF is a versatile multiplexing technology that transforms existing antibody collections into high-dimensional imaging tools without added complexity or new primary reagents. By simplifying, scaling, and adapting across modalities, OneStep-IF enables routine high-content imaging in cell biology and spatial proteomics.

Join us for this 30-minute seminar on OneStep-IF, followed by an interactive Q&A and a complimentary lunch, offering a valuable opportunity to network with colleagues and peers in the imaging community. This event is open to all King's students and staff interested in imaging - it is free, but registration is required.

Moderator

Dr Fadwa Joud, Associate Director, Innovation & Business Development, Imaging Core Facilities

Registration

Open to all King's students and staff interested in imaging, this event is free, but registration is required. Please note In-person attendance is limited, and spots will be allocated on a first-come, first-served basis. To secure your place, please register in advance.

Imaging Bites Lunch and Learn Seminar Series

This event is part of the Imaging Bites Lunch & Learn, a seminar series for King’s imaging community, designed to share knowledge and explore cutting-edge imaging topics. We invite expert speakers from academia, core facilities, and industry and each session includes a talk, interactive discussion, and complimentary lunch.

For more details and upcoming events, visit the Imaging Bites Lunch and Learn Series.