Lecture 8 (Mass Spec)

What does Mass Spec measure?

The mass of your protein or protein complex by collection mass-to-charge ratio of ions in the gas phase.

First major requirement for Mass Spec?

Converting neutral species to ions

Second major requirement for Mass Spec?

Separating the ions based on their mass-to-charge ratio

Third major requirement for Mass Spec?

Measuring the absorbance of each ion

How is the experiment performed (MS) (Step one)?

Introduce sample into the instrument

How is the experiment performed (MS) (Step two)?

using some method for ionization of your sample

How is the experiment performed (MS) (Step three)?

using a mass analyzer for separation of the ions formed

How is the experiment performed (MS) (Step four)?

detecting the ions

Mass Spec overview

1. Convert neutral atoms or molecules into a beam of positive ions

2. Separate ions on the basis of their mass-to-charge (m/z) ratio

3. Measure the relative abundance of each ion

How will the particles move in the magnetic field?

In a circular with radius dependent on mass

Name of ionization technique?

Electrospray Ionization (ESI)

Process of ESI (Step one)

the sample is forced through a capillary, in which a high voltage is applied. This creates an aerosol spray from the Taylor cone.

Process of ESI (Step two)

These droplets evaporate until the droplets reach the Rayleigh limit, where they undergo Coulomb fusion to create naked ions that move into the mass analyzer

Explanation of Rayleigh limit

A droplet tends to have a spherical shape because of the surface

tension of the liquid. If it is charged, the repulsion between ions might overcome the surface tension, leading to its explosion, forming ions.

Larger mass means?

Larger radius