Microscope calibration using laser written fluorescence

Corbett, Alexander D.; Shaw, Michael J.; Yacoot, Andrew; Jefferson, Ann; Schermelleh, Lothar; Wilson, Tony; Booth, Martin J.; Salter, Patrick S.

doi:10.1364/oe.26.021887

Cited by 37 publications

(27 citation statements)

References 22 publications

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“…The pixel size and axial image separation were determined using a specially designed femtosecond laser-etched sample slide as shown in Fig. 4 [12,11]. The calibration specimen was composed of a 1 mm thick sheet of acrylic into which was embedded a well defined array of features using a direct laser writing process detailed in [13].…”

Section: Methodsmentioning

confidence: 99%

Multi-plane remote refocusing epifluorescence microscopy to image dynamic Ca<i/>²⁺events

Lawton¹,

Saunter²,

Wilson³

et al. 2019

Biomed. Opt. Express

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Rapid imaging of multiple focal planes without sample movement may be achieved through remote refocusing, where imaging is carried out in a plane conjugate to the sample plane. The technique is ideally suited to studying the endothelial and smooth muscle cell layers of blood vessels. These are intrinsically linked through rapid communication and must be separately imaged at a sufficiently high frame rate in order to understand this biologically crucial interaction. We have designed and implemented an epifluoresence-based remote refocussing imaging system that can image each layer at up to 20fps using different dyes and excitation light for each layer, without the requirement for optically sectioning microscopy. A novel triggering system is used to activate the appropriate laser and image acquisition at each plane of interest. Using this method, we are able to achieve axial plane separations down to 15 μm, with a mean lateral stability of ≤ 0.32 μm displacement using a 60x, 1.4NA imaging objective and a 60x, 0.7NA reimaging objective. The system allows us to image and quantify endothelial cell activity and smooth muscle cell activity at a high framerate with excellent lateral and good axial resolution without requiring complex beam scanning confocal microscopes, delivering a cost effective solution for imaging two planes rapidly. We have successfully imaged and analysed Ca2+ activity of the endothelial cell layer independently of the smooth muscle layer for several minutes.

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Section: Methodsmentioning

confidence: 99%

Multi-plane remote refocusing epifluorescence microscopy to image dynamic Ca<i/>²⁺events

Lawton¹,

Saunter²,

Wilson³

et al. 2019

Biomed. Opt. Express

Self Cite

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“…Meanwhile, when using a patterned structure for remote phosphor configuration together with a non-phosphor-coated region in the perimeter area, a higher homogeneity of angular CCT and better color consistency could be achieved [18]. In addition, that the patterned sapphire substrate used in the remote phosphor is an advantage factor to get the color uniformity in the far field better than that in the conventional field [19][20][21]. Though, these aforementioned studies concentrate on the improvement of chromatic homogeneity and the lumen output of WLEDs with remote phosphor structure, they just demonstrated the results researched only on WLED packages with single LED chip and low color temperatures.…”

Section: Introductionmentioning

confidence: 99%

Study of red-emitting LaAsO4:Eu3+ phosphor for color rendering index improvement of WLEDs with dual-layer remote phosphor geometry

2020

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The remote phosphor structure is often disadvantageous in color quality but in terms of optics it is more convenient when compared to other phosphor structures such as conformal or in-cup ones. From this disadvantage, there are many studies to improve its chromatic output. In this research paper, we propose a dual-layer remote phosphor geometry for the improvement of white light-emitting diodes (WLEDs) in two parameters: color rendering index (CRI) and color quality scale (CQS). The 7700 K WLEDs are used in this study. The idea of the study is to place a red phosphor layer LaAsO4:Eu 3+ on the yellow phosphor YAG: Ce 3+ , and then find the concentration of LaAsO4:Eu 3+ in an appropriate way to achieve the highest color quality. The results showed that LaAsO4:Eu 3+ bring great benefits for enhancing CRI and CQS. In particular, the greater the concentration of LaAsO4:Eu 3+ , the greater the CRI and CQS because the portion of red lights in WLEDs increases. However, the decrease in lumen output occurs when the concentration of LaAsO4:Eu 3+ increases excessively. This is proved thanks to Mie-scattering theory and Beer-Lambert law. The results of important articles in WLEDs fabrication have greatly contributed to a higher white light quality

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“…Finally, in the works of Corbett et al and Korpelainen et al [43,44], a laser interferometer is used for measuring the vertical displacement of the equipment head and comparing it with the equipment measurements.…”

Section: Introductionmentioning

confidence: 99%

Procedure for Calibrating the Z-axis of a Confocal Microscope: Application for the Evaluation of Structured Surfaces

Wang

Caja

Gómez

et al. 2019

Sensors

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This work describes a method for the metrological characterization of structured surfaces using a confocal microscope. The proposed method is based on the calculation of texture parameters established in ISO 25178-2:2012. To ensure the traceability of these parameters, a procedure for the calibration of the Z-axis of the confocal microscope is proposed. The calculation of uncertainty associated with each parameter employs the Monte Carlo method, as well as the concept of a virtual instrument. The validity of the algorithms has been verified through the use of synthetic data provided by the National Institute of Standards and Technology (NIST) and physical standards, with minimum differences being obtained between the certified values and calculated or measured values. Finally, using the proposed method, the topography of a structured surface manufactured by laser machining is evaluated, obtaining the most used roughness parameters, as well as their measurement uncertainties and possible correlations. In general, it can be affirmed that it is possible to obtain metrologically reliable results with the proposed method.

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Microscope calibration using laser written fluorescence

Cited by 37 publications

References 22 publications

Multi-plane remote refocusing epifluorescence microscopy to image dynamic Ca<i/>²⁺events

Multi-plane remote refocusing epifluorescence microscopy to image dynamic Ca<i/>²⁺events

Study of red-emitting LaAsO4:Eu3+ phosphor for color rendering index improvement of WLEDs with dual-layer remote phosphor geometry

Procedure for Calibrating the Z-axis of a Confocal Microscope: Application for the Evaluation of Structured Surfaces

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Microscope calibration using laser written fluorescence

Cited by 37 publications

References 22 publications

Multi-plane remote refocusing epifluorescence microscopy to image dynamic Ca<i/>2+events

Multi-plane remote refocusing epifluorescence microscopy to image dynamic Ca<i/>2+events

Study of red-emitting LaAsO4:Eu3+ phosphor for color rendering index improvement of WLEDs with dual-layer remote phosphor geometry

Procedure for Calibrating the Z-axis of a Confocal Microscope: Application for the Evaluation of Structured Surfaces

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Multi-plane remote refocusing epifluorescence microscopy to image dynamic Ca<i/>²⁺events

Multi-plane remote refocusing epifluorescence microscopy to image dynamic Ca<i/>²⁺events