High throughput screen
First Claim
1. An apparatus for use with a biological cellular membrane in detecting and assaying compounds with ion channel activity, comprising:
- a well with a porous substrate disposed therein, said substrate having at least one pore with a diameter between 0.5 μ
m and 10 μ
m;
a solution perfusion channel disposed below said porous substrate and defining an inlet and an outlet for flow of fluid therethrough;
a first electrode disposed in said well;
a second electrode disposed in said solution perfusion channel in electrical communication with the first electrode through said porous substrate;
wherein said substrate is configured to permit adherence of a biological cellular membrane on an upper side thereof opposite said perfusion channel with a seal between said cellular membrane and said porous substrate having sufficient electrical resistance to facilitate patch clamping measurements across said membrane; and
further comprising a biological cellular membrane adhered only to said upper side of the porous substrate with a high resistance seal.
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Accused Products
Abstract
The present invention relates to a structure comprising a biological membrane and a porous or perforated substrate, a biological membrane, a substrate, a high throughput screen, methods for production of the structure membrane and substrate, and a method for screening a large number of test compounds in a short period. More particularly it relates to a structure comprising a biological membrane adhered to a porous or perforated substrate, a biological membrane capable of adhering with high resistance seals to a substrate such as perforated glass and the ability to form sheets having predominantly an ion channel or transporter of interest, a high throughput screen for determining the effect of test compounds on inn channel or transporter activity, methods for manufacture of the structure, membrane and substrate, and a method for monitoring ion channel or transporter activity in a membrane.
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Citations
25 Claims
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1. An apparatus for use with a biological cellular membrane in detecting and assaying compounds with ion channel activity, comprising:
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a well with a porous substrate disposed therein, said substrate having at least one pore with a diameter between 0.5 μ
m and 10 μ
m;a solution perfusion channel disposed below said porous substrate and defining an inlet and an outlet for flow of fluid therethrough; a first electrode disposed in said well; a second electrode disposed in said solution perfusion channel in electrical communication with the first electrode through said porous substrate; wherein said substrate is configured to permit adherence of a biological cellular membrane on an upper side thereof opposite said perfusion channel with a seal between said cellular membrane and said porous substrate having sufficient electrical resistance to facilitate patch clamping measurements across said membrane; and further comprising a biological cellular membrane adhered only to said upper side of the porous substrate with a high resistance seal. - View Dependent Claims (4, 5, 6, 7, 8, 9, 10, 11, 12, 13)
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2. An apparatus for use with a biological cellular membrane in detecting and assaying compounds with ion channel activity, comprising:
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a well with a porous substrate disposed therein, said substrate having at least one pore with a diameter between 0.5 μ
m and 10 μ
m;a solution perfusion channel disposed below said porous substrate and defining an inlet and an outlet for flow of fluid therethrough; a first electrode disposed in said well; a second electrode disposed in said solution perfusion channel in electrical communication with the first electrode through said porous substrate; wherein said substrate is configured to permit adherence of a biological cellular membrane on an upper side thereof opposite said perfusion channel with a seal between said cellular membrane and said porous substrate having sufficient electrical resistance to facilitate patch clamping measurements across said membrane; and further comprising a voltage-clamp amplifier connected between said electrodes and configured for patch clamping measurements. - View Dependent Claims (3)
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14. An apparatus for use with a biological cellular membrane in detecting and assaying compounds with ion channel activity, comprising:
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a multi-well plate defining a plurality of wells, each with a laser perforated substrate disposed therein, said substrate being perforated with plural pores in each said well having a pore diameter of 1 μ
m-2 μ
m, and said substrate being configured to permit adherence of a biological cellular membrane thereto with a seal having an electrical resistance sufficient to facilitate patch clamping measurements thereacross;a solution perfusion channel disposed below said porous substrate with a differential pressure applied across said substrate to create lower pressure below the substrate to facilitate positioning of cells on said pores said perfusion channel having an inlet and an outlet; at least a first electrode disposed on a moveable recording head for sequentially positioning in said plurality of wells; a second electrode disposed in said solution perfusion channel in electrical communication with the first electrode through said porous substrate; a voltage-clamp amplifier connected between said first and second electrodes for patch clamping measurements. - View Dependent Claims (15, 16, 17, 18, 19, 20, 21)
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22. A high throughput apparatus for use with a biological cellular membrane in detecting and assaying compounds with ion channel activity, comprising:
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a multi-well plate defining a plurality of wells; a perforated plastic substrate disposed at a bottom of said wells, said substrate being perforated with plural pores in each said well, said pores having a diameter of 1 μ
m-2 μ
m;a solution perfusion channel disposed below said porous substrate with a differential pressure applied across said substrate to create a lower pressure below the substrate to facilitate positioning of cells on said pores, said perfusion channel having an inlet and an outlet; plural first electrodes disposed on a moveable recording head for sequentially positioning in said plurality of wells; at least a second electrode disposed below said porous substrate in electrical communication with said first electrodes through said porous substrate; and a voltage-clamp amplifier connected between said electrodes for patch clamping measurements. - View Dependent Claims (23, 24, 25)
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Specification