Biological and Biomaterials preparation, image, and characterization laboratory


Zeiss spinning disc confocal

Hitachi S-570 LaB6 SEM

Hitachi S-900 Field-emission SEM

Preparatory equipment

Colloidal metals & conjugates

Instrument & training fees

Practical Microscopy course

Microscopy links

Making and conjugating colloidal metals

Colloidal metal particles are commonly used to label features of interest for electron microscopy. Electron-dense metals are used, as in the transmission EM they block the beam electrons, producing a black shape in the image, and in the SEM, they show as brighter spots on the labeled objects. For SEM, back-scattered (BSE) electron detectors can be used to positively identify the colloidal metals, since heavy metals are a better source of BSEs, and therefore form brighter spots in the image.

The difficulty to date with colloidal metal labeling is that one black (or bright) spot looks like any other, so that the only way to do multiple labeling for EM was to use colloidal particles of different sizes. This method is limited in usefulness, however, as only 2 or maybe 3 labels could be found of sufficienly distinct size to distinguish the labels, and the large particles could easily cause steric interference and prevent binding of other labeled moities. The way around this problem is to produce small labels (small relative to the targets) of different compositions or shapes. The labels can then be easily distinguished by their composition using e.g., Electron Energy-Loss Spectroscopy (EELS) in the TEM, or by their shapes using either TEM or SEM.

These colloidal metal particles are then conjugated to antibodies and other molecules to produce labels for various targets. Conjugation takes place at a pH where the ligand molecule is electrically neutral. For an protein, this is a pH slightly alkaline to the protein's pI. Cell surface receptors, epitopes of connective tissues, or most anything else for which an appropriate antibody or ligand can be found can be targeted by such a conjugated colloid-ligand complex.

The BBPIC makes colloidal metals of different compositions and sizes, and conjugates them to proteins and other ligands, and this is a service for use by any group that needs it. Making the colloids and doing the conjugations can be done by BBPIC staff, or investigators may be trained in the procedures.

Some colloidal metal particles produced in the BBPIC for conjugating to proteins and other ligands
Metal Shape Size
Gold round 1 to several hundred nm
gold facetted (triangles, hexagons, pentagons, trapezoids) 50 to 500 nm
gold oblong (teardrops, dumbbells) 50 to 500 nm
silver round 5 to 10 nm
palladium umbonate ("popcorn") 18 nm
palladium facetted (triangles, cubes, pentagons, hexagons) 10 to 15 nm
palladium round 30 to 40 nm
gold-coated platinum round 5 to 20 nm
different sizes of colloidal gold particles
Image by Irawati Kandela, Dept. Pharmaceutical Sciences, Sch. of Pharmacy, UW-Madison, & Ralph Albrecht lab, Dept. Animal Sciences.
Colloidal metals image gallery
27 Minute Quick Time Video on making Colloidal Gold
Please contact Ralph Albrecht or Joe Heintz for further information.