FRSC 365 Phase contrast

What is phase contrast microscopy?

A contrast-enhancing technique used to increase visibility of transparent specimens.

What types of specimens are typically used in phase contrast microscopy?

Living cells, microorganisms, thin tissue slices, glass fragments, and sub-cellular particles.

Why can transparent objects be difficult to see under a microscope?

Because the intensity of the sample may be very similar to the non-scattered light, making them hard to distinguish.

What is zero order light in phase contrast microscopy?

Non-scattered, undeviated light that passes through the specimen.

What happens to light rays when they pass through a transparent specimen?

Some light is not scattered (zero order light) while other rays are diffracted.

What is the goal of using a phase shifter in phase contrast microscopy?

To create destructive interference between diffracted and undeviated rays, increasing specimen contrast.

What is a phase plate?

A component installed at the objective back focal plane that alters the phase of light to enhance contrast.

How does a phase shifter affect the light in phase contrast microscopy?

It causes the undeviated light and diffracted light to become out-of-phase, enhancing contrast.

What is the result of constructive interference in phase contrast microscopy?

It forms a bright image on a darker background, known as negative phase contrast.

What is the purpose of an apodized phase plate?

To reduce unwanted halo effects around the specimen.

What is the significance of aligning the condenser annulus with the phase shifter?

It is crucial for proper phase contrast imaging.

What are some limitations of phase contrast microscopy?

Halos and optical artifacts can obscure details, and the phase annuli can limit the numerical aperture, reducing resolution.

What is the difference between positive and negative phase contrast?

Positive phase contrast results in a dark specimen against a light background, while negative phase contrast results in a bright specimen against a dark background.

What is the role of a thin absorbing layer on the phase shifter?

It balances the intensity of undeviated and diffracted light, enhancing contrast.

What does 'Ph' refer to in microscopy objectives?

Phase contrast objective.

What is the purpose of using monochromatic light in phase contrast microscopy?

To reduce halo effects caused by optical artifacts.

What is the function of the phase telescope or Bertrand lens?

It is used to view the alignment of the phase shifter and condenser annulus.

What is the effect of a phase shifter that is thicker than the rest of the plate?

It slows undeviated light, causing it to arrive in-phase with diffracted light, leading to bright phase contrast.

What does 'S' represent in the context of phase contrast microscopy?

'S' represents the surround wave or undeviated light.

What does 'D' represent in the context of phase contrast microscopy?

'D' represents the diffracted wave.

What does 'P' represent in the context of phase contrast microscopy?

'P' represents the particle wave resulting from interference between S and D.