Please always reserve time on microscopes in iLabs and remove reservation if you need to cancel. Both microscopes in this core are biannually serviced and have been updated to the latest software that works for each unit. Microscopes areonly as good as the person using them, so please ask if you are having issues.
Having a reasonable understanding of how microscopes work will make it easy to assess if something isn’t right with your sample or the microscope.
Microscopy: (General knowledge and review)
Properties of Light: (light source)
Fluorophore selection: (This site will explain your excitation and emission requirements for imaging)
Nikon Eclipse Ti2-E Inverted Motorized Research Microscope System
CellSens software: (https://www.olympus-lifescience.com/en/software/cellsens/?gclid=EAIaIQobChMI-qLug-3h6wIVT77ACh0BvA3eEAAYASAAEgJc8_D_BwE) Basic version
This unit utilizes MetaMorph Software. Cameras: (https://www.photometrics.com/wp-content/uploads/2020/03/MicroPublisher-3.3-5.0-Datasheet.pdf)
Filter cubes for Nikon
The anatomy of an objective is important to getting the best image possible from all the hard work of preparing your sample. The following applies to OIL objectives.
- NEVER add oil to an objective UNLESS is says oil on the objective barrel.
- Independent of the microscope being inverted or upright, the slide should be situated so the objective is looking through the coverslip and not the slide.
- Cover glass thickness matters for resolution, this is found after the infinity symbol (6)
- It is a good practice to make sure that the retraction stopper isn’t frozen in place by excess, old oil. If it is stiff, you could break the coverslip. (With lens paper over lens, gently press until you feel the objective give under your finger). (5)
- After working with the OIL objective, please, using lens paper, gently clean the surface. Remove the objective and return to its holder and place in left top drawer. CLEAN your slide as well.
- Immersion oil-Type NHV, designed for inverted scopes is a higher viscosity oil designed to remain in place between coverslip and objective. Immersion oil-Type FF is a nearly, fluorescence free oil, meant to reduce background fluorescence. This core has both lens paper and glass cleaning wipes located in metal cabinet.
- The higher the numerical (NA), the higher the resolving power. Using Nikon objectives from above:
- 20X = NA (0.50)
- 40X = NA (0.75)
- 60X = NA (1.40)
- The higher the NA, the shallower the depth of focus (DOF) DOF = The DOF is greatest on the lowest power objective (4X/10X). Each time you switch to a higher power (20X/40X/oil objectives [60X/100X]), the DOF is reduced. Therefore, a smaller part of the specimen is in focus at higher power. The amount of light transmitted to your eye is greatest at the low power WD = working distance of objective (for the image above, is 0.14mm) is the distance between the lens of the objective and the surface of your coverslip.