Ny DHM-studie

"Vår" cancerforskare Robert L. Judson-Torres m kollegor har genomfört en gedigen studie om Digital Holografisk Mikroskopi (DHM). Man går igenom tekniken och berättar om alla dess fördelar och användningsområden. Studien är en bra genomgång om hur tekniken utvecklats från enklare funktioner till att idag klara alltmer avancerade observationer.Man lyfter de ev tillkortakommanden som fortfarande finns till att bli ett "standardinstrument" för den breda forskningen. Där kommer man även med svar på hur lösa det, riktade till instrumenttillverkarna. En kul detalj är att Robert tar upp utvecklingen från tidig upptäckt om förlagan till DHM som då gav upphovsmännen ett Nobelpris. Som  många av er vet har bloggen ett gott öga till Robert och ser honom som en kommande Nobelpristagare med sin nydanande forskning inom hudcancer.

Denna studie tillsammans med vår andre HoloMonitorfrälstes , den ungerskfödde forskaren Robert Horvath, studie som publicerades i Nature alldeles nyligen är guld värt för att möjliggöra att tekniken ska få sitt breda genombrott. HoloMonitor i Nature.

Dessa 2 Robertar drar onekligen en lans för tekniken i allmänhet och för HoloMonitor i synnerhet.

Ägna gärna en stund åt att läsa igenom den nu aktuella studien som publicerades igår 2 Augusti.

Quantitative Phase Imaging: Recent Advances and Expanding Potential in Biomedicine

Abstract

Quantitative phase imaging (QPI) is a label-free, wide-field microscopy approach with significant opportunities for biomedical applications. QPI uses the natural phase shift of light as it passes through a transparent object, such as a mammalian cell, to quantify biomass distribution and spatial and temporal changes in biomass. Reported in cell studies more than 60 years ago, ongoing advances in QPI hardware and software are leading to numerous applications in biology, with a dramatic expansion in utility over the past two decades. Today, investigations of cell size, morphology, behavior, cellular viscoelasticity, drug efficacy, biomass accumulation and turnover, and transport mechanics are supporting studies of development, physiology, neural activity, cancer, and additional physiological processes and diseases. Here, we review the field of QPI in biology starting with underlying principles, followed by a discussion of technical approaches currently available or being developed, and end with an examination of the breadth of applications in use or under development. We comment on strengths and shortcomings for the deployment of QPI in key biomedical contexts and conclude with emerging challenges and opportunities based on combining QPI with other methodologies that expand the scope and utility of QPI even further.

Digital Holography

Digital holography directly descends from interferometry and also captures the interference between a reference and sample beam. However, unlike interferometry, digital holography does not typically require mechanical scanning of the resulting interference fringes. In digital holography, the interferogram is captured with a digital camera placed at a known distance in front of the image plane. This interferogram is analyzed using diffraction theory to reconstruct the complex object wavefront, including the phase shift and intensity modulation of light passing through the sample. Digital holography emerged from the establishment of holography by Gabor for which he won the Nobel prize in 1971. Gabor’s work demonstrated that light from a point source interfering with secondary waves from light scattered by an object produces a negative photograph of a three-dimensional (3D) image. However, a conjugate image is also superimposed on the reconstructed image, resulting in ambiguity due to the presence of this twin image. It was later shown that use of an off-axis reference beam can separate the real and conjugate image. Marine plankton provided an early application of live cells imaged using holography in a chamber with close proximity to a photographic plate.

I nedanstående stycke klipper jag in områden Robert L Judson-Torres identifierar som högst användbara för tekniken. Ni får läsa resp rubriks innehåll själva.

Advances in Quantitative Biology

As QPI approaches have advanced, so too have QPI applications. One advantage of QPI is that it is label-free. Therefore, QPI can study cell behavior with minimal impact, a leveraged feature in a number of biological applications. As summarized above, there are also a number of additional label-free microscopy approaches, including the more widely used methods of phase contrast and DIC microscopy. The primary advantage of QPI over these other approaches, however, is that, in contrast to phase contrast or DIC microscopy, the data contained in each pixel of a QPI image are a quantitative measure of the phase delay of light as it passes through that portion of a sample. Measurement of this phase delay can utilize any of the approaches already discussed above. Once captured, analysis of this phase data can provide quantitative insights into numerous biological processes and systems. Here we summarize key advances in the application of QPI to quantitative studies in biology, ranging from applications that quantify the behavior of individual cells to emerging opportunities in clinical diagnostics

QPI Applications Using Measurements of Cell Mass or Growth Rate

Applications of QPI to Studies of Cell Growth and Associated Biological Processes

Applications of QPI to Studies of Immune Cell Behavior

Applications of QPI to Measure Neuron Behavior

Applications of QPI in Measuring the Physical Structure of a Cell

Applications of QPI in Studies of Intracellular Transport

Applications of QPI to Cell Migration Assays

Applications of QPI for Measuring Biophysical Cell Properties

QPI Applications in Screening and Drug Sensitivity Measuremen

QPI Morphological Applications in Diagnostics

 

Conclusions and Perspective

QPI is an approach with a long history. However, the last two decades have seen great leaps in both the abilities and applications of QPI. The rapid recent development of QPI is from impressive advances in image processing capabilities enabled by digitalization and increasing computational power . 

This development and application of computational tools has substantially increased the utility and power of QPI in its application to biomedicine and permitted the development and commercialization of prebuilt and user-friendly QPI platforms. Consequently, recent years have witnessed a surging interest in QPI, coupled to a dramatic increase in QPI enabled publications and discoveries.

 

This marked expansion of QPI applications is also being fueled by leveraging machine learning approaches and is increasingly impacting areas that are beginning to include disease diagnoses and measurements of biological state transitions. While exciting, this recent and rapid adoption of QPI platforms and associated published studies has also highlighted the dearth of standardization tools and practices beyond the adaptation of polystyrene beads as phase standards. Developing and circulating such tools will be critical for reproducible studies and validation of future QPI-based diagnostics and other applications.

 

Current areas of QPI utility include studies of cell size and its regulation, cellular diagnostics and screens, and biomechanics and biophysics. One key strength of QPI approaches includes label-free classification of key cellular behaviors such as programmed cell death pathways, differentiation, cell cycle progression, and immunological responses. Assessing these behaviors in the context of changes in biomass density, morphology, transport, and viscoelastic properties provides a deeper understanding of adaptations during cell or organismal life. A second key strength is the ability to study single cells or individual cell clusters over long periods of time. 

 

As techniques in single cell profiling continue development resulting in increasing reports on molecularly distinct subpopulations of cells, QPI provides a platform for assessing distinct phenotypes and behaviors within these heterogeneous populations. Further development of multimodal approaches will be critical for merging the observations made using single cell molecular profiling with QPI single cell phenotyping.

 

Finally, although there have been a large number of studies pointing toward clinical utility of QPI, this approach is ready for more robust validation and testing with clinical samples. As a label-free approach that can quantify multiple physiologically relevant parameters describing the behavior of living cells, QPI is well positioned to work with clinical samples.

 

QPI therefore has the potential to enable a wide range of clinical applications in functional and diagnostic medicine, both as an addition to current approaches that rely on staining and as an independent ex vivo approach. Further work is therefore needed to build on the demonstrated capabilities of QPI to translate this technology to clinical utility and ultimately to improve the standard of patient care.

 

Min kommentar

Med denna imponerande studie har Robert L. Judson-Torres m kollegor banat väg för tekniken att nå fler,många fler användare inom en massa forskningsområden som inbegriper rubrikerna ovan.

Robert nämner PHI och HoloMonitor i texten (som sig bör). 

Nu håller vi tummarna att studien sprids brett till (den ack så konservativa) forskarvärlden.

 

Tillägg 4/8

Studien har nu lagts upp i ett mer exklusivt format. Länk (PDF)

                                           Mvh the99

 

Bonus

Mexikansk (!) doktorsavhandling om hudcancer som enbart bygger på DHM.

 

Título :

MICROSCOPÍA HOLOGRÁFICA DIGITAL PARA LA CARACTERIZACIÓN DE VARIABLES FÍSICAS EN MUESTRAS DE CÉLULAS DE PIEL

 

Resumen o descripción:

"Se implementó un sistema de DHM para la medición del índice de refracción y espesor en una muestra deshidratada de la línea celular de melanoma A375, acompañado de un novedoso modelo matemático para desacoplar estas variables. Adicionalmente, se configuró un sistema de DHM para obtener mapas de fase desenvueltos ópticamente. Con este arreglo, se realizaron mediciones de espesor y longitud en las capas epidérmicas córnea, espinosa, basal y sus células individuales correspondientes a muestras de una biopsia con cáncer de piel tipo SCC de bajo grado."

 

Doktoranden refererar till studier utförda av Robert L. Judson-Torres och hänvisar till studier utförda med HoloMonitor.

Klicka på PDF längst ner på sidan så kan ni läsa hela studien (på spanska).