Multifocal Imaging Research Articles

Multi-dimensional resolution of elementary Ca 2+ signals by simultaneous multi-focal imaging

Elementary events such as puffs and sparks are cytosolic microdomains of Ca 2+ from which cellular Ca 2+ signals are constructed. Because of the tight localization and fast kinetics of elementary events, imaging studies have been hindered by instrumental limitations of confocal and deconvolution fluorescence microscopy which necessitate compromises between spatial and temporal resolution. Here, we describe a novel, yet simple ‘multi-focal’ fluorescence microscopy system that employs three high-speed cameras focused at different axial depths to enable 4-dimensional imaging with millisecond resolution. We demonstrate the utility of this system for studies of puffs in Xenopus oocytes by mapping the axial distribution of puff sites, by obtaining measurements of puff amplitudes undistorted by focus error, and by deriving deblurred images that reveal novel sub-micron jumps of Ca 2+ release sites.

Retinotopic distribution of chromatic responses in human primary visual cortex

In non-human primates at least three anatomically and functionally distinct channels convey signals from the retina to the primary visual cortex (V1). Two of these channels, the parvocellular and the koniocellular, are sensitive to chromatic contrasts and form the basis of color vision. In humans, common phylogenetic history with other primates and psychophysical experiments suggest identical retinocortical mechanisms but separate evaluation of the distinct anatomical channels has been difficult because signals are already combined in V1. We studied the spatial distribution of activation to chromatic stimuli along the two opponent chromatic axes in human V1 with multifocal functional magnetic resonance imaging. The signal strength was quantified from three experiments with stimuli up to 20 degrees eccentricity. The hypothesis was that, although the parvo- and koniocellular signals are mixed in V1, distinct distributions of signal strength would be evident. We found that whereas different conditions activated the same areas of cortex, indicating that they have identical magnification factors, the responses to red/green stimulation were stronger close to the fovea whereas the blue/yellow responses were much less diminished with increasing eccentricity. Both chromatic axes showed saturating contrast response functions. Our measure directly from human V1 is in line with earlier psychophysical studies suggesting relatively stronger parvocellular channel representation close to the fovea, and more uniform distribution of the koniocellular and achromatic channels. In addition, our study presents a way to rapidly quantify retinotopic signal transmission in distinct retinocortical pathways of individual subjects.

An integrated approach to fast and informative morphological vouchering of nematodes for applications in molecular barcoding

Molecular surveys of meiofaunal diversity face some interesting methodological challenges when it comes to interstitial nematodes from soils and sediments. Morphology-based surveys are greatly limited in processing speed, while barcoding approaches for nematodes are hampered by difficulties of matching sequence data with traditional taxonomy. Intermediate technology is needed to bridge the gap between both approaches. An example of such technology is video capture and editing microscopy, which consists of the recording of taxonomically informative multifocal series of microscopy images as digital video clips. The integration of multifocal imaging with sequence analysis of the D2D3 region of large subunit (LSU) rDNA is illustrated here in the context of a combined morphological and barcode sequencing survey of marine nematodes from Baja California and California. The resulting video clips and sequence data are made available online in the database NemATOL (http://nematol.unh.edu/). Analyses of 37 barcoded nematodes suggest that these represent at least 32 species, none of which matches available D2D3 sequences in public databases. The recorded multifocal vouchers allowed us to identify most specimens to genus, and will be used to match specimens with subsequent species identifications and descriptions of preserved specimens. Like molecular barcodes, multifocal voucher archives are part of a wider effort at structuring and changing the process of biodiversity discovery. We argue that data-rich surveys and phylogenetic tools for analysis of barcode sequences are an essential component of the exploration of phyla with a high fraction of undiscovered species. Our methods are also directly applicable to other meiofauna such as for example gastrotrichs and tardigrades.

Multifocal Three-Dimensional Imaging with Optical Lattice Excitation

Extract HTML view is not available for this content. However, as you have access to this content, a full PDF is available via the ‘Save PDF’ action button. Extended abstract of a paper presented at Microscopy and Microanalysis 2005 in Honolulu, Hawaii, USA, July 31--August 4, 2005

Laser Beam Splitting by Diffractive Optics

Recent advances in diffractive optics theory and technology have made it possible to design programmable, multichannel optical systems based on diffraction gratings, lenslet arrays and digital holography algorithms. Spot array generation, multiple and multifocal imaging, matched filtering, laser beam mode selection and simultaneous contour shaping are among the most promising applications of diffractive beam splitting.

Time-decorrelated multifocal micromachining and trapping

Temporally decorrelated multifocal arrays eliminate spatial interference between adjacent foci and allow multifocal imaging with the diffraction-limited resolution of a single focus, even for foci spaced by less than the focal diameter. In this paper, we demonstrate a high-efficiency cascaded-beamsplitter array for producing temporally decorrelated beamlets. These beamlets are used to produce a multifocal microscope with which we have demonstrated two-photon fluorescence imaging, multifocal micromachining of optical waveguides, and multifocal optical trapping.

Time-decorrelated multifocal array for multiphoton microscopy and micromachining

We demonstrate a temporally decorrelated, multifocal multiphoton microscope. Using an etalon, we split the 800-nm light from either an ultrashort-pulsed Ti:Al (2)O (3) oscillator or a Ti:Al (2)O (3) regenerative amplifier into an array of beamlets that are delayed with respect to one another in time. The collimated beams overlap at slightly different input angles at the entrance pupil of a 1.25-numerical aperture oil-immersion objective to produce an array of foci that are temporally decorrelated at the focal plane of the objective. The temporal decorrelation eliminates any interference among the foci and permits multifocal multiphoton imaging with the resolution of single-point illumination.

Diagnosis and therapy of pediatric and juvenile hematogenous osteomyelitis

Haematogen osteomyelitis is mostly found in children and adolescents. In western Europe acute haematogen osteomyelitis (AHOM) is a rare disease. This is the cause why AHOM is often diagnosed with delay. The treatment usually is an antibiotic medication and/or surgical interventions. Uncharacteristic pain of extremities in children should always consider the diagnosis of acute osteomyelitis. Investigation should include conventional X-rays, ultrasounds or MRI to prevent the spreading of infection. In cases of multifocal infection radionuclide imaging should be undergone. Differential diagnosis should always include malignant tumor. If under treatment of antibiotics the clinical signs of illness do not decrease within 24 h surgery with fenestration of the involved bone, debridement and local application of antibiotics is indicated. In unusual cases or in cases with clinical signs of AHOM but no bacteria specification a malignant tumor has to be excluded.

Multifocal enhancing magnetic resonance imaging lesions following cranial irradiation

Radiation necrosis is a delayed complication of cranial irradiation, typically presenting as a single intracerebral mass that is radiographically indistinguishable from recurrent tumor. We describe 6 patients with a distinct radiographic syndrome of multifocal enhancing lesions on magnetic resonance images, and their variable clinical courses: some fluctuating, some spontaneously resolving, and some demonstrating fulminant progression to frank necrosis.