Super-resolution microscopy can identify specific protein distribution patterns in platelets incubated with cancer cells

Abstract

Super-resolution imaging of P-selectin in platelets together with dictionary learning allow specifically activated platelets to be identified in an automatic objective manner.

Platelet-Cancer Interplay: Molecular Mechanisms and New Therapeutic Avenues

“…However, the concept of co-clustering of proteins in distinct granules and differential granule release was challenged by several studies using quantitative enzyme-linked immunosorbent assay (ELISA), confocal immunofluorescence microscopy and proteomic approaches ( 60 – 64 ), which did not observe any functional pattern. Recently, it has been proposed that Stimulated Emission Depletion (STED) imaging can be applied to study platelet granule content and protein clustering in a more precise manner ( 65 , 66 ). Whether depending on the time course and the activating stimuli, platelets can enhance or inhibit tumor angiogenesis by selective release of pro- or anti-angiogenic factors is an intriguing question, which needs to be addressed in an in vivo context.…”

Quantitative Optical Diffraction Tomography Imaging of Mouse Platelets

“…RI index scales; Pink: 1.3406-1.3525, blue: 1.3525-1.3621, green: 1.3622-1.3735, red: 1.3736-1.3822. **P ≤ 0.01, ****P ≤ 0.0001. structure and function of platelets or platelet proteins during activation processes or disorders has recently been produced (Poulter et al, 2015;Westmoreland et al, 2016;Knight et al, 2017;Bergstrand et al, 2019;Khan and Pike, 2020). With fluorescence based imaging, the protein of interest often requires the samples to have a fluorescent tag -either genetically modified or by immuno-labeling, to provide the contrast required for visualization.…”

“…Imaging based techniques are rising in popularity with the advancement of super-resolution imaging and the amount of quantitative information concerning the functioning of the cells that can be derived from such experiments. The field of platelet biology has taken advantage of these tools and a vast array of new and exciting information relating to the changes in the internal structure and function of platelets or platelet proteins during activation processes or disorders has recently been produced ( Poulter et al, 2015 ; Westmoreland et al, 2016 ; Knight et al, 2017 ; Bergstrand et al, 2019 ; Khan and Pike, 2020 ). With fluorescence based imaging, the protein of interest often requires the samples to have a fluorescent tag – either genetically modified or by immuno-labeling, to provide the contrast required for visualization.…”

“…The limited volume of the blood obtained for isolation of platelets severely restricts the number and type of assays that can be performed with mouse blood. Super-resolution imaging techniques have recently enabled the study of structural changes in the protein distribution or cytoskeleton changes of platelets in disease models (Poulter et al, 2015;Bergstrand et al, 2019;Khan and Pike, 2020), However, it is important to note that with improvements in resolution, the post-isolation processing which entails fixation, permeabilization and labeling, can often lead to artifacts if not carefully controlled (Schnell et al, 2012;Stanly et al, 2016;Pereira et al, 2019;Khan and Pike, 2020).…”

Stimulated emission depletion microscopy for biological imaging in four dimensions: A review

“…The typical STED resolution ranges from 30 to 70 nm, as recorded with the commercial STED systems, such as Leica TCS SP8 STED (Gemmink et al, 2018; Masch et al, 2018; Thompson et al, 2017; C. Wang et al, 2019; Yang et al, 2020) or Abberior's STED systems (Bergstrand et al, 2019; Carravilla et al, 2019; Kamper, Ta, Jensen, Hell, & Jakobs, 2018; Soliman, Göttfert, Rosewich, Thoms, & Gärtner, 2018; Stephan, Roesch, Riedel, & Jakobs, 2019), as well as several custom‐built STED systems. Abberior STEDYCON can achieve resolution <40 nm, when using 640 nm as excitation beam and 775 nm as STED beam, 100× 1.45 NA objective.…”