Success stories

Human brain tumors

"PLA is a standard in our lab representing a state of the art quantitative protein technology that has furthered our research on human brain tumors above and beyond classical protein techniques."

Markus Bredel, MD, PhD, Northwestern University, Chicago
Cell focal adhesions

"The availability of the Duolink system could not have come at a better time for my investigation of two novel proteins found to be present in cell focal adhesions. My research has established that both proteins co-localise using double immunofluroesecent labelling. However, there are many disadvantages to using immunofluorescent labelling for this particular application such as limits in the resolution of optical microscopy in addition to fluorescent bleed-through and cross-talk. As a result of these problems, complex analysis is required to statistically determine whether there is true co-localisation, and can be laborious and time consuming. Therefore, the in situ proximity ligation assay by Duolink is advantageous for this particular study in that it will confirm the results of the co-localisation study without the afore mentioned disadvantages. So far, I have found the system to be simple to use and have obtained positive results."

Gethin Owen (PhD), Periodontal Biology Laboratory, Oral, Biol. Med Sci., Faculty of Dentistry, University of British Columbia, Vancouver, Canada
Visualize endogenous interactions

"By using proximity ligation assays, PLA, we were able to confirm data from a 2-hybrid screen and from co-immunoprecipitation experiments that demonstrated a dynamic interaction between a membrane receptor and a transcription factor. Some of the features we love about PLA are: we are able to visualize endogenous interactions using just a small number of cells- which is good, even crucial, when primary or difficult to growth cells are used; furthermore we were able to detect protein-protein interactions and to define the cellular localization in which the interactions occurred at the same time; finally because the signal produced by PLA can be amplified, the method is exquisitely sensitive. The technique also allows quantification (which everybody, including study sections and reviewers evaluating papers submitted for publication, loves). If good primary antibodies are available, PLA is a powerful and easy to use technique."

Roxana Pincheira. PhD University of California San Francisco
Wound healing and cancer metastasis

"Duolink in situ PLA is a sensitive tool that provides new insights into the temporal and spatial changes in protein-protein profile. It has become an indispensible technique in our research on wound healing and cancer metastasis."

Andrew Tan, PhD, Nanyang Technological University, Singapore
Protein-protein interactions

"The Duolink kit allowed us to confirm in cells a receptor complex we had detected by immuno-precipitation analysis.  This technique may ultimately prove to be equivalent to FRET but with the significant benefit of detecting endogenous protein-protein interactions."

David A. Cheresh, Ph.D., Professor and Vice Chair of Pathology, University of California, San Diego, Moores Cancer Center
Visualization of cell signalling in cancer

"In situ PLA has allowed us to visualize the interaction of endogenous mdm2 and p53 following various drug treatments, and to evaluate inhibition of protein-protein interactions with small molecules. The technology allows visualization of the cellular compartment (nucleus, cytoplasm, etc) in which protein-protein interactions take place, and thus can offer a significant advantage over conventional (lysis-based) immunoprecipitation techniques. As PLA requires very few cells, it also helps conserve limited biological samples and reagents."

Mark Wade. PhD, Gene Expression Laboratory (Wahl Lab), The Salk Institute, San Diego
Post translational modifications detected in FFPE liver tissue

"Fluorescence colocalization in paraffin-embedded liver tissue is complicated by the lipofuscin-associated autofluorescence in the specimens.  The increase in signal-to-noise attained with the application of the proximity ligation assay allowed us to easily extract out of the background the acquisition of true colocalized signals validating the observation that post-translational nitrotyrosine modification of the mitochondrial F1F0 Complex V ATPase occurs during endotoxin-induced nitrooxidative stress in cattle."

Ted. H. Elsasser, PhD, United States Department of Agriculture, Beltsville, MD
Subcellular localization of protein interactions and modifications

"The Duolink kit has become a standard method in our lab for the detection and quantification of protein interactions. In combination with good primary antibodies, the method allows for sensitive, yet fast and easy detection and quantification of endogenous interactions. For experiments with transfected cells, it offers the advantage over co-immunoprecipitations that the transfected cell population can be analyzed separately from the untransfected cell population. By using phospho-specific antibodies, we have also successfully used PLA to quantify phosphorylation events. We also like that we get information about the subcellular localization where these interactions and phosphorylation events occur."

Susanne Vetterkind, PhD, Boston University, Boston
Novel biomarker research studies

“The low sample consumption and overall immunoassay performance of Proseek has enabled us to access precious and limited biobanked plasma samples, a commonly recurring problem with standard technologies which use 50-100 fold more material. We are now able to perform novel biomarker research studies with the goal of improving early detection of breast and colorectal cancer.”

Jan Stenvang, PhD, University of Copenhagen, Denmark
Protein quantification in precious samples

“The Proseek immunoassay allows detection and quantification of low abundant proteins in biological fluids using very small sample amounts. The system is user-friendly and allows simultaneous analysis of many samples in a 96-well format. These particular properties render it a very useful tool for protein quantification in very precious samples, such as human sera, suitable for high throughput screenings. To the best of our knowledge, Proseek largely exceeds the performance of conventional immunoassays. We think that this assay can be successfully exploited for protein expression profiles in complex biological samples and biomarker discovery.”

Renata Grifantini, Research Head, Externautics Spa, Italy
Forensic medicine application of PLA

The age estimation of injuries remains a challenge to forensic pathologists. By using Duolink and primary antibodies against proteins that interact in the coagulation cascade and in the complement system we could observe that some combinations of primary antibodies produce temporally distinct reactivity. Hence, we found that the combination P-Selectin and PSGL-1 produces a distinct reactivity in the early phase after injury, whereas other combinations of factors in the cascade react somewhat later. We could also observe that the reactivity was absent in undamaged tissue, and when the interval between injury and death was too short or too long. This all-or-none response of a particular protein interaction is attractive since it means that a time window for positive reactivity can be established for each antibody combination. And the distinct staining pattern provided by the Duolink technique allows for an evaluation that is easy even for untrained observers to carry out. Using a set of antibody combinations against protein interactions with different time windows, age determination of injuries can be improved and help police investigations in e.g. obscure homicide cases.

Henrik Druid, Professor of Forensic Medicine, Karolinska Institutet, Sweden