US 5,585,275 A · Pilot apparatus for peptide synthesis and screening

Abstract

Method and apparatus for simple and rapid preparation of reusable, addressable surface-immobilized arrays of biomolecules (libraries) used for screening for interaction with any biologically significant target. A special plate having on or in its surface a plurality of discreet functionalized substrate areas, typically in arrays of 10.times.10 to 400.times.400, is provided for chemical synthesis or bonding thereon of desired families of biomolecules (e.g. peptides, DNA, RNA, oligosaccharides). In the case of peptides, such as hexapeptides, the resulting permanently hexapeptide-loaded plate is a reusable Addressable Synthetic Peptide Combinatorial Library (ASPCL), in which 1 to 3 (typically two) of the positions in the sequence are uniquely identified by the address location. The preferred plate embodiment employs an HPMP wink of porous polyolefin removably received in holes in the plate. A unique multi-slot block assembly is used to prepare the ASPCLs. The wink carrier plate is also employed with a vacuum block system to assist in washing, deprotection, and probing. In library applications, for example determining peptides which bind to functional proteins (enzymes, receptors, antibodies), the substrate-bound peptides are assembled with several positions consisting of uniformly distributed equimolar mixtures of residues, and 2 separated or sequential positions uniquely identified by their spatial location on the substrate array, the "address". Following identification of the known residues giving the greatest affinity for the arrayed positions in the sequence, optimal binding for the complete peptide sequence is determined by an iterative process replacing formerly mixed positions with known AAs at unique addresses.

Claims17

Dependent claims

A biological screening assembly comprising in operative combination:

a) a one piece, inert substrate carrier plate having an array of discrete, individually addressable areas for receiving a substrate material, said substrate material chemically functionalized for assembly of biopolymers thereon;
b) a substrate material selected from the group consisting of:
i) an acid chloride activated surface;
ii) a particulate material;
iii) a functionalized polyolefin disc;
and
iv) a functionalized porous polyolefin disc;
wherein said substrate material in at least some of said discrete, individually addressable areas is coated with a hydrophilic polar multifunctionalized polysaccharide chain covalently coupled to said substrate material;
wherein said hydrophilic polar multi-functionalized polysaccharide chain is covalently coupled at multiple points to said substrate material spaced along the length of each polysaccharide chain, wherein said covalent bonds are present in an amount sufficient to minimize loss of said polysaccharide chain from said support upon exposure to trifluoroacetic acid and to minimize the effect of hydrolysis in said polysaccharide chain during peptide synthesis deprotection, to form a thin film three dimensional functionalized matrix layer above said substrate in an open, freely permeable configuration for the covalent tethering of ligands thereon, and for access thereinto of target molecules for affinity binding to said ligands which are tetherable to said thin film three dimensional functionalized matrix by covalent bonds for affinity screening of target molecules, in which at least one of said substrate material is secured to each of said discrete, individually addressable areas;
and each of said substrate material is wholly contained within the boundary of each discrete individually addressable area.

7 more elements

A biological screening system comprising in operative combination:

a) an inert support plate having an array of a plurality of discrete, individually addressable areas for receiving a substrate material;
b) substrate material chemically functionalized for assembly of biopolymers thereon secured to each of said addressable areas in which said substrate material is wholly contained within the boundary of each discrete, individually addressable area;
c) an elastomeric barrier gasket sheet overlying said inert support plate having an array of holes therethrough aligned with said array of discrete, individually addressable areas for access to said substrate material, said holes independent and isolated from one another;
and
d) an apertured block disposed on said elastomeric barrier gasket, said apertured block having a plurality of slots passing therethrough, each of said slots oriented to provide a common well for simultaneous access of reagents and biomonomer solutions for the synthesis of biopolymers to a plurality of said substrate materials in said array.

2 more elements

Dependencies are inferred from the claim text.

Pilot apparatus for peptide synthesis and screening

Life of this patent

Pilot apparatus for peptide synthesis and screening

Life of this patent

Abstract

Claims17