Who with Whom? Visualising Protein Relationships with Naveni® Plex from Navinci

Releasing the brakes on the immune system - this is the mechanism behind so-called immune checkpoint inhibitors, which are already being used for numerous types of tumours, such as malignant skin cancer, lung cancer or head and neck tumours. This special form of cancer immunotherapy targets molecular checkpoints that regulate the immune response under normal conditions and prevent healthy cells from being damaged by excessive defence processes. Tumour cells use precisely these checkpoints to evade immune surveillance. This is where checkpoint inhibitors come in: They ‘release the brakes’ and enable the immune system to specifically recognise and fight tumour cells [1]. Key interactions between surface molecules on tumour and immune cells play a central role here.

As treatment with immune checkpoint inhibitors is only effective in some patients, functional biomarkers are urgently needed in order to better predict the response to cancer therapies and provide more targeted treatment. A deeper understanding of the complex protein relationships in the tumour microenvironment is essential for this. This is precisely where the Naveni® Plex from Navinci Diagnostics comes in: With this innovative multiplex solution based on the in situ proximity ligation assay (isPLA), several crucial protein interactions can be visualised simultaneously - precisely, spatially resolved and functionally relevant.

These topics await you:

1) Three in one - these protein relationships are uncovered by Naveni® Plex

2) How it works - the Naveni® in situ proximity ligation technology

3) Sensitivity, Specificity, Resolution - why you should choose Naveni® Plex

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Three in one - these protein relationships are uncovered by Naveni® Plex

Understanding cellular signalling pathways is crucial for the targeted development of immune checkpoint therapies. Naveni® Plex enables the simultaneous detection of central immuno-oncological biomarkers directly in tissue samples - and thus contributes to a more precise assessment of therapy effectiveness and better patient stratification [2]. The isPLA visualises these crucial protein relationships:

PD-1/PD-L1 interactions: show activated immune checkpoint signalling pathways
CD8/MHC-I interactions: mark the start of central T cell activation processes
CD3 detection: identifies T cells as a central component of the immune response

While CD3 serves as a general marker for T cells, the CD8/MHC-I interaction is a central step in the activation of cytotoxic T cells - a prerequisite for the recognition and elimination of tumour cells. The PD-1/PD-L1 interaction, on the other hand, has an inhibitory effect and leads to the exhaustion of immune cells - a known mechanism of immune escape. The simultaneous detection of PD-1/PD-L1 and CD8/MHC-I makes it possible to investigate how immune escape and T-cell activation are connected - and how tumours specifically block immune attacks [3].

**How it works - the Naveni® in situ proximity ligation technology**

The kit is based on the innovative in situ proximity ligation assay (isPLA) technology, which utilises specially developed primary antibodies, so-called navenibodies. These are each conjugated with proprietary oligonucleotide arms (Fig. 1). A specific pair of navenibodies is available for each target molecule - either against two different proteins to detect interactions (PD1/PD-L1 and CD8/MHC-I), or against two different epitopes of the same protein to detect individual target proteins in a highly specific manner (CD3). Only when both navenibodies bind to their target structures and are in close proximity to each other is a rolling circle amplification (RCA) reaction triggered, which generates a strong, specific fluorescence signal (Fig. 1). A separate fluorophore is used for each target, which enables simultaneous detection of several interactions in one sample: Cy3 for PD1/PD-L1, Cy5 for CD8/MHC-I and FITC for CD3 [4].

NaveniPlex_Figure1

Figure 1: The Naveni® in situ proximity ligation technology. Step 1: Navenibodies bind to their target proteins. Step 2: Navenibodies that are spatially close to each other are connected via a circularised DNA ‘bridge’. Step 3: The connecting DNA is amplified and detected by multiple fluorescence-labelled oligonucleotides [5].

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Sensitivity, Specificity, Resolution - why you should choose Naveni® Plex

As the first multiplex isPLA, the Naveni® Plex marks another milestone in the history of innovation at Navinci Diagnostics. The Swedish biotechnology company has established itself as a pioneer in the development of in situ proximity ligation assay technology and impresses with excellent solutions that help researchers worldwide to comprehensively analyse protein interactions in a tissue context [2]. Naveni® Plex is characterised by exceptionally high specificity, sensitivity and resolution and can be easily combined with other spatial analysis methods. Thanks to the separate provision of the Navenibodies, the assay can be flexibly configured: Users can select different combinations and create customised panels for their research questions [3].

The advantages of Naveni® Plex at a glance:

Multiplex isPLA for functional proteomics: Simultaneous detection of multiple protein interactions in the tumour-immune context - beyond pure expression data to functional significance
Outstanding sensitivity and specificity: Dual antibody recognition plus signal amplification ensure high-precision detection - even for challenging targets in multiplex format
Ready-to-use and easy to integrate: Ready-to-use kits with carefully validated reagents - compatible with established in situ workflows and other spatial technologies