Oil immersion

In light microscopy, oil immersion is a technique used to increase the resolving power of a microscope. This is achieved by immersing both the objective lens and the specimen in a transparent oil of high refractive index, thereby increasing the numerical aperture of the objective lens. Without oil, light waves reflect off the slide specimen through the glass cover slip, through the air, and into the microscope lens (see the colored figure to the right). Unless a wave comes out at a 90-degree angle, it bends when it hits a new substance, the amount of bend depending on the angle. This distorts the image. Air has a very different index of refraction from glass, making for a larger bend compared to oil, which has an index more similar to glass. Specially manufactured oil can have nearly exactly the same refractive index as glass, making an oil immersed lens nearly as effective as having entirely glass to the sample (which would be impractical). Immersion oils are transparent oils that have specific optical and viscosity characteristics necessary for use in microscopy. Typical oils used have an index of refraction of around 1.515. An oil immersion objective is an objective lens specially designed to be used in this way. Many condensers also give optimal resolution when the condenser lens is immersed in oil. Lenses reconstruct the light scattered by an object. To successfully achieve this end, ideally, all the diffraction orders have to be collected. This is related to the opening angle of the lens and its refractive index. The resolution of a microscope is defined as the minimum separation needed between two objects under examination in order for the microscope to discern them as separate objects. This minimum distance is labelled δ. If two objects are separated by a distance shorter than δ, they will appear as a single object in the microscope. A measure of the resolving power, R.P., of a lens is given by its numerical aperture, NA: where λ is the wavelength of light.