Ichrome mle

Manufactured by Toptica

Sourced in United States

The IChrome MLE is a compact, diode-based laser source that generates coherent light in the green spectral range. It provides a stable output power and a narrow linewidth, making it suitable for various applications requiring high-quality green laser light.

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Lab products found in correlation

Ixon ultra, by Oxford Instruments (3 mentions) Linear polarizer, by Thorlabs (2 mentions) F 15 cm lenses, by Thorlabs (2 mentions) Ff02 617 73, by IDEX Corporation (2 mentions) Ixon ultra 897, by Oxford Instruments (2 mentions) Ff01 520 35 25, by IDEX Corporation (1 mentions) Spectralis hra oct, by Heidelberg Engineering (1 mentions) Ff01 676 29 25, by IDEX Corporation (1 mentions) Dfc365fx, by Leica (1 mentions) Ff01 676 37 25, by IDEX Corporation (1 mentions) Spectra x, by Lumencor (1 mentions) Orca flash 4, by Hamamatsu Photonics (1 mentions) Nis elements software, by Nikon (1 mentions) Ix81 microscope, by Olympus (1 mentions) Axio imager z2, by Zeiss (1 mentions) Orca flash4.0 v3, by Hamamatsu Photonics (1 mentions) Umplfln 10 w, by Olympus (1 mentions) 4.2 plus, by Oxford Instruments (1 mentions)

Spelling variants of this product

Four-line iChrome MLE laser (2 citations) IChrome MLE-L multilaser engine (2 citations)

13 protocols using ichrome mle

Volumetric Telomere Imaging in Cells

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For telomere imaging in fixed cells, the 4f system consisted of two f=15 cm lenses (Thorlabs), a linear polarizer (Thorlabs) to filter out the light that is polarized in the unmodulated direction of the LC-SLM, a 1920 × 1080 pixel LC-SLM (PLUTO-VIS, Holoeye) and a mirror for beam-steering. A sCMOS camera (Prime95B, Photometrics) was used to record the data. The sample was illuminated with 561 nm fiber-coupled laser-light source (iChrome MLE, Toptica). The excitation light was reflected up through the microscope objective by a multibandpass dichroic filter (TRF89902-EM - ET - 405/488/561/647 nm Laser Quad Band Set, Chroma). Emission light was filtered by the same dichroic and also filtered by another 617 nm band pass filter (FF02-617/73 Semrock).
For volumetric telomere tracking in live cells, images were recorded with an EMCCD camera (Andor iXON), exposure time 100 ms and EM-gain 170. The sample was illuminated at ≈2 kW/cm² with 561 nm light from a fiber-coupled laser (iChrome MLE, Toptica). All movies were recorded for 50 s (500 frames).

Nehme E., Freedman D., Gordon R., Ferdman B., Weiss L.E., Alalouf O., Naor T., Orange R., Michaeli T, & Shechtman Y. (2020). DeepSTORM3D: dense 3D localization microscopy and PSF design by deep learning. Nature methods, 17(7), 734-740.

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Volumetric Telomere Imaging in Cells

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Adaptive Optics Imaging of Retinal Cells

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Mice were imaged with a custom adaptive optics scanning light ophthalmoscope (AOSLO), using near-infrared light (796∆17 nm, 200-500 µW, super luminescent diode: S790-G-I-15, Superlum, Ireland) [2, 17] . Phase-contrast imaging referred to in the context of this paper, was achieved by purposefully displacing the detector axially to a plane conjugate to the highly reflective RPE/choroid complex, to enable detection of forward and multiply scattered light from translucent cells, as detailed in our recent publication [5] . In a subset of experiments for confirmation of immune cell types, fluorescence was simultaneously imaged using 488 nm excitation and 520Δ35 emission for GFP, and 640 nm excitation and 676Δ29 emission for Alexa Fluor 647 (excitation laser diode: iChrome MLE, Toptica Photonics, Farmington, New York, USA; emission filters: FF01-520/35-25 and FF01-676/29-25, Semrock, Rochester, New York, USA). Mice also underwent imaging with HRA+OCT Spectralis (Heidelberg Engineering, Germany).

Joseph A., Chu C.J., Feng G., Dholakia K., & Schallek J. (2020). Label-free imaging of immune cell dynamics in the living retina using adaptive optics.

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Cryo-SOFI Imaging for Fluorescence Microscopy

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We modified a commercial cryo-FM system (Cryo CLEM; Leica) equipped with a 50× 0.9-NA objective lens (Cryo CLEM Objective HCX PL APO 50×/0.9; Leica) for cryo-SOFI imaging by coupling lasers (iChrome MLE; Toptica) into the microscope body for fluorophore excitation and photoswitching. Laser light after the single-mode fiber was collimated using an achromatic lens with 19-mm focal length to achieve an illuminated area of ∼40-µm diameter in the object plane. A schematic illustration of additionally required hardware is in SI Appendix, Fig. S1. For all biological examples presented here, data were acquired using 488-nm laser illumination and the standard GFP filter cube of the microscope system (excitation: 470/40; dichroic: 495 low pass; emission: 525/50). Overview images where recorded using a standard CCD camera (DFC365 FX; Leica). Cryo-SOFI data were recorded with an electron multiplying CCD (EMCCD) camera (iXon Ultra 897; Andor) and additional 2× magnification to reach an overall magnification of 100× for matching the larger pixel size of the EMCCD camera. Typical camera settings for cryo-SOFI data acquisition were 50-ms integration time per frame (at a rate of 20 frames per second) for a time series of 2,000 images at an EM gain of 20–200.

Moser F., Pražák V., Mordhorst V., Andrade D.M., Baker L.A., Hagen C., Grünewald K, & Kaufmann R. (2019). Cryo-SOFI enabling low-dose super-resolution correlative light and electron cryo-microscopy. Proceedings of the National Academy of Sciences of the United States of America, 116(11), 4804-4809.

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Single-Molecule Live Cell Imaging

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Live cell single-molecule tracking was performed at room temperature on a custom-built TIRF microscope built around the Rapid Automated Modular Microscope (RAMM) System (ASI Imaging). Laser lines of 488 and 561 nm provided by a multi-laser engine were used to activate GFP and TMR/mCherry, respectively (iChrome MLE, Toptica). Laser beams were collimated at the fibre output and focused (×100 oil-immersion objective, NA 1.4, Olympus) onto the sample at an angle allowing for highly inclined thin illumination^{38 (link),39}. Fluorescence emission was filtered by a dichroic mirror and notch filter (ZT405/488/561rpc & ZET405/488/561NF, Chroma) and then projected onto an EMCCD camera (iXon Ultra, 512 × 512 pixels, Andor). The pixel size was 96 nm in the magnified image. Transmission illumination was provided by an LED source and condenser (ASI Imaging). Sample position and focus were controlled with a motorised piezo stage, a z-motor objective mount, and autofocus system (MS-2000, PZ-2000FT, CRISP, ASI Imaging). All video data were collected at room temperature (20–22 °C), with a 33 ms frame rate. Moderate bleaching of the sample was applied to be able to track single molecules, as described below.

Rassam P., Long K.R., Kaminska R., Williams D.J., Papadakos G., Baumann C.G, & Kleanthous C. (2018). Intermembrane crosstalk drives inner-membrane protein organization in Escherichia coli. Nature Communications, 9, 1082.

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Dual-color TIRF Microscopy of Yeast

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The yeast dual-color data were acquired on a microscope with a commercial laser box (iChrome MLE, Toptica) with 405 nm, 561 nm and 640 nm lasers and a 640 nm booster laser (Toptica), which were coupled via single mode. The output of the fiber was collimated, focused on the back focal plane of the TIRF objective (60×/NA 1.49, Nikon), and adjusted for epi illumination. The emitted fluorescence was laterally constricted by a slit, split by a dichroic mirror (640LP, ZT640rdc, Chroma), filtered by the respective bandpass filters (transmitted/AF647: 676/37, FF01-676/37-25, Semrock; reflected/mMaple: 600/60, NC458462, Chroma), and imaged on two parts of the EMCCD camera (iXON Ultra, Andor). The focus was stabilized as described for the system above. Raw data were acquired with a 30 ms exposure time. The images acquired in the two channels were merged using a transformation that was determined using images of beads that are fluorescent in both channels (TetraSpeck).

Wu Y.L., Hoess P., Tschanz A., Matti U., Mund M, & Ries J. (2022). Maximum-likelihood model fitting for quantitative analysis of SMLM data. Nature Methods, 20(1), 139-148.

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Multimodal Microscopy Imaging Setup

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A 100X oil-immersion objective (UAPON100XOTIRF) an EMCCD camera (iXon Ultra 897, Andor) coupled to an inverted microscope (IX83, Olympus), fitted with both epifluorescence and TIRF illumination optics, were used for all imaging experiments. An LED light source (SPECTRA X, Lumencor) was used for epifluorescence illumination and a laser (iChrome MLE, Toptica Photonics) for TIRF illumination.

Grupi A., Shapira Z., Degani-Katzav N., Yudovich S., & Weiss S. (2021). Point-localized and site-specific membrane potential optical recording by engineered single fluorescent nanodiscs.

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Super-resolution Imaging with CoDiM Module

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Super-resolution imaging was performed with a commercial CoDiM (Caron et al., 2014 (link); Fallet et al., 2014 (link)). The BioAxial super-resolution module (CODIM100, BioAxial) is an add-on integrated to the Nikon confocal system previously described. The CODIM module acts as a powerful beam shaper generating local structured illumination. A sCMOS camera plugged at the back port of the microscope (Orca Flash 4.0, Hamamatsu Photonics) is used for the detection generating individual micro-images for each scanning point containing independent information. The set of all micro-images obtained from the scan procedure are processed and reconstructed by CODIM algorithm (Nesterov, 2005 (link)) to generate a super-resolved image. In both confocal and super-resolution modalities, a 60x 1.49NA Oil immersion Nikon Plan Apo TIRF objective was used to focus the laser beam and collect of the emitted fluorescence. The 488 and 561 excitation wavelengths of a multi laser engine (iChrome MLE, Toptica Photonics Inc.) are used for fluorescence excitation. The confocal image captures were performed using Nikon Nis-Elements software (Nikon Instruments Europe). The laser power was properly chosen to be out of the saturation regime. Confocal and super-resolution montages were subsequently built in ImageJ (NIH).

Garita-Hernandez M., Guibbal L., Toualbi L., Routet F., Chaffiol A., Winckler C., Harinquet M., Robert C., Fouquet S., Bellow S., Sahel J.A., Goureau O., Duebel J, & Dalkara D. (2018). Optogenetic Light Sensors in Human Retinal Organoids. Frontiers in Neuroscience, 12, 789.

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Imaging Primary Hippocampal Neuron Cultures

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Rat primary hippocampal cultures were imaged on DIV 16–18. The external solution for bath perfusion contained (in mM): 145 NaCl, 2.5 KCl, 10 HEPES, 2 CaCl₂, 1 MgCl₂, and 10 glucose (pH 7.3). Excitation by 488- and 561-nm lasers (iChrome MLE, Toptica Photonics) for GFP and Red Fluorescent Protein (RFP) was directed through a manual TIRF input to an Olympus IX81 microscope. We used a ×100 Olympus objective (UAPON 100x O TIRF, NA 1.49) for all recordings on bulk culture primary hippocampal neurons. Nonetheless, we did not image in TIRF mode because bleaching was too severe in this condition. Fluorescence intensities in response to 488- and 561-nm excitation were recorded sequentially with 20-ms exposure time and 1 × 1 binning on a Prime 95B CMOS camera (1200x1200 pixels, 11 µm pixel size; Photometrics). GFP and RFP emission were split with a H 560 LPXR superflat beamsplitter (AHF, Germany), and recorded on the same frame after passing through ET525/50 m (GFP) and ET620/60 m (RFP) filters (both Chroma). Emission filters were mounted within an Optosplit II Bypass (Cairn Research). Exposures were timed to precede the −80 mV voltage steps. Images were recorded with MicroManager and analyzed with Fiji as described above.

Hao Y., Toulmé E., König B., Rosenmund C, & Plested A.J. (2023). Targeted sensors for glutamatergic neurotransmission. eLife, 12, e84029.

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Live Cell Single-Molecule Tracking via TIRF

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Live cell photoactivated single-molecule tracking was performed on a custom-built total internal reflection fluorescence (TIRF) microscope built around the Rapid Automated Modular Microscope (RAMM) System (ASI Imaging) as previously described [1, (link)24] (link). PAmCherry activation was controlled by a 405 nm laser and excited with 561 nm. All lasers were provided by a multi-laser engine (iChrome MLE, Toptica). At the fiber output, the laser beams were collimated and focused (100x oil immersion objective, NA 1.4, Olympus) onto the sample under an angle allowing for highly inclined thin illumination (Tokunaga et al., 2008) . Fluorescence emission was filtered by a dichroic mirror and filter (ZT405/488/561rpc & ZET405/488/561NF, Chroma). PAmCherry emission was projected onto an EMCCD camera (iXon Ultra, 512x512 pixels, Andor). The pixel size was 96 nm. Transmission illumination was provided by an LED source and condenser (ASI Imaging and Olympus). Sample position and focus were controlled with a motorized piezo stage, a z-motor objective mount, and autofocus system (MS-2000, PZ-2000FT, CRISP, ASI Imaging). Movies of 10,000 frames at 23ºC were acquired under continuous 561 nm laser excitation at 250 W/cm 2 with an exposure time of 15 ms. Camera readout was 0.48 ms giving frame intervals of 15.48 ms.

Kaja E., Vijande D., Kowalczyk J., Michalak M., Gapiński J., Kobras C., Rolfe P, & Stracy M. (2024). Comparing Mfd- and UvrD-dependent models of transcription coupled DNA repair in live Escherichia coli using single-molecule tracking. DNA repair, 137.

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