Columns

Depending on the application, immunoaffinity matrices can be packed into open columns, such as screening columns (Fisher Scientific, Pittsburgh, PA), and a variety of glass columns, such as those provided by Amersham Pharmacia Biotechnology, Bio-Rad, or other vendors. The column material must be compatible with your analysis because either plastic or glass can cause problems with low concentrations of some analytes. Column dimensions are dependent

Table 1

Some Examples of Commercially Available Gels Used for the Immobilization of Antibody

Table 1

Some Examples of Commercially Available Gels Used for the Immobilization of Antibody

Function group

Group to

Matrix

on the beads

be coupled

Products0

Cross-linked agarose

-CHO

-NH2

Aminolink coupling gel (P)

diaminopropylamine

-COOH

Immobilized diaminodipropylamine (P)

cyanate

-NH2

CNBr activated matrices (S) CNBr-activated Sepharose 4B (A)

Imidazolyl carbamate

-NH2

carbonyldiimidazole affinity resins (S) Reacti-gel (P)

N-hydroxysuccinimide

-NH2

HNS activated matrices (S) Affi-gel 10 (B), Affi-gel 15 (B)

iodoacetyl

-SH

Ultralink iodoacetyl (P)

hydrazide

-CHO

Agarose Adipic Acid Hydrazide (A)

Bis-acrylamide/azlactone

-NH2, -SH, -OH,

Ultralink biosupport medium (P)

Acrylic

N-hydroxysuccinimide

-NH2

Affi- Prep 10 (B)

Pressure stable polymer

hydrazide

-CHO

Affi-Prep Hz (B)

"The letter in parenthesis indicates the abbreviation of the commercial sources. A: Amersham Pharmacia Biotechnology; B: Bio-Rad; P: Pierce; S: Sigma. This is not an exhaustive list, but serves to provide examples of the most commonly used activated supports.

"The letter in parenthesis indicates the abbreviation of the commercial sources. A: Amersham Pharmacia Biotechnology; B: Bio-Rad; P: Pierce; S: Sigma. This is not an exhaustive list, but serves to provide examples of the most commonly used activated supports.

on specific applications; we routinely use plastic columns having a 5-mL capacity and pack them with 1 mL of support. Normally we pack the columns using gravity flow; but depending upon the support either pressure or vacuum assisted packing techniques might be appropriate. An optimum flow rate (approx 0.5 mL/min) permits equilibration between the immobilized antibody and the analyte.

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