I’ve recently obtained a Nikon Microphot-FXA BODY to convert into a microphotographic do it all. One reason I choose this project is my long affiliation with Nikon instruments and Nikon cameras. Although a short, 79pg operators manual is available, I’m having difficulty obtaining any technical manuals. Does anyone have access to one or know where it could be obtained? Issues are what objectives and light sources are compatible, etc. This will be refitted with digital still and video cameras in place of the old film cameras. Any help greatly appreciated. I know it originally had CF objectives ..read more

Hi all, I’m looking for anti GFP fluorescent nanobodies conjugated with Alexa 647 (or equivalent) for immunostaining of samples with GFP in order to do dSTORM imaging. Does anyone know of any commercially available fluorescent nanobodies that target GFP? So far I have found the NanoTag FluoTag options and ChromoTek GFP-boosters. 1 post - 1 participant Read full topic ..read more

Hi everyone, Would anyone willing to share some feedback or experience with their fluorescent lamp ? We are looking to buy a new one for a basic widefield (no live application) and are open to any suggestion in this choice ! If you have some references to share, feel free. Thanks Julien 1 post - 1 participant Read full topic ..read more

Hi everybody, We are interested in dyes that perform well with 2P imaging. We have large structures stained with primary Ab and secondary Ab conjugated to fluorophores. Alexa488 works fine for us, but we would like to extend the palette of colours / multiplex. Has someone experience with this? Thanks and best regards, Laurent. 1 post - 1 participant Read full topic ..read more

Hello all. I have a Leica DMi8 TIRF microscope with a TIRF lens, 100x/1.47. The camera pixel size is 6.5um. I’d like to use it for dSTORM, and needs a 60x TIRF lens to reach ~100nm/pixel. Unfortunately, Leica has discontinued their 63x TIRF lens. I recently got an Olympus 60x/1.49 TIRF lens (APON60XOTIRF). I’m wondering if it’s possible to use an adapter, M25 to RMS, in order to use Olympus 60x on the DMi8. Do you see any disadvantage of this mix and match strategy? Thank you. 4 posts - 3 participants Read full topic ..read more

Hello, wondering if anyone may have suggestions to a problem we’re having with our PhenoCycler-Fusion (CODEX) from Akoya Biosciences. We’ve been running it well for several months, but recently we’ve run into issues where certain antibodies that we’ve conjugated ourselves only stain the edge of tissue. During antibody validation, we did not see the same staining patterns via IHC (on the same tissue, serial sections). We’ve tried a variety of things to solve this issue including changing out all buffers, trying multiple variations of antigen retrieval, staining and incubation, etc., but it hasn ..read more

The PUMA with its improved lower collector Köhler system can deliver full Köhler illumination with objectives as low as x2 - as shown here: I am unaware of any other microscope that can provide full Köhler illumination with full Abbe condenser down to as low as x2 so if you know of one I would be grateful if anyone can tell me (outside custom built instruments on optical benches). Thanks. Field illumination curvature in both images above is exaggerated by the wide angle camera used to take these pictures via afocal projection through the eyepiece of both scopes. If you would like to see more ..read more

Hello everyone, I do multiplexe immunofluorescence and I use zeiss axio imager for imaging my slides. I noticed that when I set the focus on DAPI channel, the focus in other channels (FITC, cy3 and cy5) is not as sharp as in DAPI channel, especially in the red spectrum. I believe this is something known and one way to work around it is to set a Z offset to correct for this small difference between channels. I would be grateful to know what is the best way to set the correct Z offset between channels? I for now use the Z stack option and I just set the right focus of DAPI channel, set it as the ..read more

Hello all. I was trying to calculate the illumination area in fluorescence microscopy to estimate the irradiance. Can you help me confirm if the following is correct? In point scanning confocal, we overfill the objective BFP to create a diffraction-limited illumination spot on the sample. The illumination area = pi * square (0.61*lamda / NA)/(4 * ln(2)). The lamda is the Ex wavelength, and NA is the objective numerical aperture, and 4 * ln(2) is the gaussian beam estimation. In the case of Yokogawa spinning disk confocal, can I still use the above equation to calculate the single illumination ..read more

Continuing the discussion from Guide to Microscopy about Cellular Ploidy Variation: Good morning My name is Matheus, I’m starting out in the area of image analysis and I would like to develop an analysis and identification of polyploid cells in histological slides stained in HE and also with fluorescence. I would like to know if there was any guide that highlights how to identify and characterize polyploid and multinucleated cells in slides observed using conventional or fluorescence microscopy? Thanks a lot for the help. 1 post - 1 participant Read full topic ..read more