pulled Protocols

Category: Microinjection Protocols: Microinjection Equipment

This protocol describes an easy method for calibrating micropipette tips that have been pulled in the laboratory. It is essential to estimate the internal diameter of the pulled micropipette tip when adjusting parameters for a new puller or new type of glass tubing. A tip diameter of ~0.3 µm is optimal for the microinjection of mammalian cells in culture (e.g., CHO, PtK1, and COS-7). A 10% increase in diameter increases the delivery rate by more than 30% and can cause cell damage. - [Read Calibration of Micropipette Tips Protocols]

Category: Microinjection Protocols: Microinjection Equipment

This protocol contains methods for pulling microinjection needles using two different models of pipette pullers. The advantage of pulling needles in the laboratory is that a variety of different needle types can be pulled, depending on the samples and cells being injected. An added advantage is cost; once a pipette puller has been purchased, boxes of glass capillaries are inexpensive compared to premade microinjection needles. - [Read Preparation (Pulling) of Needles for Gene Delivery by Microinjection Protocol]

Category: Microinjection Protocols: Microinjection of Proteins Protocols

his protocol provides methods for the preparation of protein samples and for loading them into pulled microinjection pipettes. Stock solutions of proteins are thawed, diluted (if desired), centrifuged at high speed to remove aggregates, and kept on ice until loading. Loading into micropipettes can be done using either a "front-loading" or a "backfilling" procedure. - [Read Preparation and Loading of Protein Samples for Microinjection Protocol]