Blood

Single cell protein interactomes by the proximity network assay

F Filip Karlsson (1Pixelgen Technologies, Stockholm, Sweden) M Michele Simonetti (1Pixelgen Technologies, Stockholm, Sweden) C Christina Galonska (1Pixelgen Technologies, Stockholm, Sweden) M Max Karlsson (1Pixelgen Technologies, Stockholm, Sweden) H Hanna van Ooijen (1Pixelgen Technologies, Stockholm, Sweden) T Tomasz Kallas (1Pixelgen Technologies, Stockholm, Sweden) D Divya Thiagrajan (1Pixelgen Technologies, Stockholm, Sweden) M Maud Schweitzer (1Pixelgen Technologies, Stockholm, Sweden) L Ludvig Larsson P Pouria Tajvar (1Pixelgen Technologies, Stockholm, Sweden) J Johan Dahlberg (1Pixelgen Technologies, Stockholm, Sweden) V Vincent van Hoef (1Pixelgen Technologies, Stockholm, Sweden) F Florian De Temmerman (2BioLizard nv, Ghent, Belgium) L Louise Leijonancker (1Pixelgen Technologies, Stockholm, Sweden) V Vanessa Thrombin (1Pixelgen Technologies, Stockholm, Sweden) S Sylvain Geny (1Pixelgen Technologies, Stockholm, Sweden) R Rikard Forlin (3Karolinska Institutet, Department of Women's and Children's Health, Solna, Sweden) E Erika Negrini (3Karolinska Institutet, Department of Women's and Children's Health, Solna, Sweden) S Stefan Petkov (5Pixelgen Technologies, Burlington, Canada) L Lovisa Franzén J Jessica Bunz (1Pixelgen Technologies, Stockholm, Sweden) C Christine Moge (1Pixelgen Technologies, Stockholm, Sweden) H Henrik Everberg (1Pixelgen Technologies, Stockholm, Sweden) P Petter Brodin A Alvaro Martinez Barrio (1Pixelgen Technologies, Stockholm, Sweden) S Simon Fredriksson (5Pixelgen Technologies, Burlington, Canada)

Abstract

Abstract Cellular function depends on dynamic interactions and nanoscale spatial organisation of proteins. While transcriptomic and proteomic methods have enabled single-cell profiling, scalable technologies allowing high-resolution analysis of protein interactions at omics-scale are lacking. Here we present the Proximity Network Assay (PNA), a DNA-based method for constructing three-dimensional nanoscale maps of 155 proteins in single cells without the use of optics. PNA employs barcoded antibodies and in situ rolling circle amplification to generate >40,000 spatial nodes per cell, which are linked through proximity-dependent gap-fill ligation and decoded by DNA sequencing, forming single cell Proximity Networks. At an estimated spatial resolution of ~50 nm, PNA captures single-cell protein abundance, self-clustering, and colocalization, validating established cell membrane protein interactions. We illustrate how PNA can be used to gain insights into the molecular mechanisms of cell function through protein interactions in hematological oncology, CAR-T cell therapies, and autoimmune disease.

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Article Details

Journal Blood
Volume / Issue Vol. 146, Issue Supplement 1
Published November 03, 2025
Pages 6570-6570
ISSN 0006-4971
Publisher Elsevier BV

Journal Info

Blood

Elsevier BV
ISSN: 0006-4971 Health Sciences

Authors (26)

F

Filip Karlsson

1Pixelgen Technologies, Stockholm, Sweden

M

Michele Simonetti

1Pixelgen Technologies, Stockholm, Sweden

C

Christina Galonska

1Pixelgen Technologies, Stockholm, Sweden

M

Max Karlsson

1Pixelgen Technologies, Stockholm, Sweden

H

Hanna van Ooijen

1Pixelgen Technologies, Stockholm, Sweden

T

Tomasz Kallas

1Pixelgen Technologies, Stockholm, Sweden

D

Divya Thiagrajan

1Pixelgen Technologies, Stockholm, Sweden

M

Maud Schweitzer

1Pixelgen Technologies, Stockholm, Sweden

L

Ludvig Larsson

P

Pouria Tajvar

1Pixelgen Technologies, Stockholm, Sweden

J

Johan Dahlberg

1Pixelgen Technologies, Stockholm, Sweden

V

Vincent van Hoef

1Pixelgen Technologies, Stockholm, Sweden

F

Florian De Temmerman

2BioLizard nv, Ghent, Belgium

L

Louise Leijonancker

1Pixelgen Technologies, Stockholm, Sweden

V

Vanessa Thrombin

1Pixelgen Technologies, Stockholm, Sweden

S

Sylvain Geny

1Pixelgen Technologies, Stockholm, Sweden

R

Rikard Forlin

3Karolinska Institutet, Department of Women's and Children's Health, Solna, Sweden

E

Erika Negrini

3Karolinska Institutet, Department of Women's and Children's Health, Solna, Sweden

S

Stefan Petkov

5Pixelgen Technologies, Burlington, Canada

L

Lovisa Franzén

J

Jessica Bunz

1Pixelgen Technologies, Stockholm, Sweden

C

Christine Moge

1Pixelgen Technologies, Stockholm, Sweden

H

Henrik Everberg

1Pixelgen Technologies, Stockholm, Sweden

P

Petter Brodin

A

Alvaro Martinez Barrio

1Pixelgen Technologies, Stockholm, Sweden

S

Simon Fredriksson

5Pixelgen Technologies, Burlington, Canada

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