The present invention relates generally to methods for defining regions of differing porosity of a nitrocellulose film on a substrate. Specifically, the present invention is related to forming patterns on nitrocellulose slides using a suitable solvent vapor to define a lower porosity region as oppose to high porosity region for the purpose of defining regions of differing porosity of a nitrocellulose film on a substrate. More specifically, the present invention is related to forming patterns on nitrocellulose slides using a suitable solvent vapor to define a lower porosity region as oppose to a high porosity region for purpose of defining regions of differing porosity of a nitrocellulose film on a substrate using an XZY robot and flat hypodermic needle.
Nitrocellulose is a common material used for binding of proteins for biochemical assays such as antibody/antigen reactions. Nitrocellulose membranes currently are the main support matrix for “rapid test” products such as over the counter urine tests (U.S. Pat. No. 6,818,455 & U.S. Pat. No. 5,602,040) as well as a variety of blood tests. Theses rapid test products are readily usable by an unskilled person and which preferably merely requires that some portion of the product is contacted with the sample (e.g. a urine stream in the case of a pregnancy or ovulation test) and thereafter no further actions are required by the user before an analytical result can be observed. Typically, the analytical result should be observable within a matter of minutes following sample application, e.g. ten minutes or less.
The high protein binding capacity and reliable “wicking” ability of the membranes has secured their use in the market for many years. With the advent of microarray techniques there has been interest in providing nitrocellulose films on glass slide substrates [1,2]. Most commonly such films have a high degree of porosity, are typically white, and are usually provided as defined regions on a glass slide. Since the films are porous, defined regions are necessary to perform multiple experiments on 1 slide without cross-contamination. Examples of such slides are available from GE Healthcare (Whatman FAST Slides), Grace Bio Labs (ONCYTE Slides) and more recently by Schott. In most cases the nitrocellulose film is applied to the glass using a spin casting method. Following casting of the porous film, the nitrocellulose is removed in unwanted areas to define regions or “pads”. Alternatively a transparent nitrocellulose film (U.S. Pat. No. 6,861,251) is available from GenTel Biosciences (PATH Slides). This is a non-porous film and therefore defined regions are not necessary. In most cases slides are mounted in a frame that facilitates the processing of multiple assays at one time such as in U.S. Pat. No. 7,063,979.
Although the technology has been fairly mature as to casting porous films, the removal process of nitrocellulose in unwanted areas to define regions or “pads” needs vast improvements. Specifically, it is unresolved that an effective, fast, efficient and economical method to remove nitrocellulose in unwanted area to define region of pads is needed to reduce the costs of preparing nitrocellulose film on a substrate wherein such substrate is capable of performing multiple immunoassay experiments.
It is an object of this invention to provide a method to effectively and efficiently define regions of differing porosity of a nitrocellulose film on a substrate.
It is an object of this invention to provide a method to effectively and efficiently pattern nitrocellulose slide on a substrate.
It is a further an object of this invention to provide a method of pattern nitrocellulose slide in a cost effective manner.
It is further an object of this invention to provide an apparatus to pattern nitrocellulose slide in a cost effective manner
It is further an object of this invention to provide an apparatus that efficiently defines regions of differing porosity of a nitrocellulose film on a substrate.
The present invention relates to the production of defined regions of differing porosity of a nitrocellulose(NC)-based film on a substrate. Starting with a substrate which has a nitrocellulose-based film of uniform porosity, areas of the film are exposed to a flow of a suitable solvent vapor such that the film in the exposed area is dissolved in the solvent vapor and upon removal of the solvent vapor will dry as a film of lower porosity than the original film. The lower porosity region is then sufficient to separate (usually using a frame with a rubber seal) multiple experiments that are performed simultaneously on the substrate.
In one embodiment, a method of forming a pattern on a nitrocellulose film on a substrate is provided comprising: providing a nitrocellulose based film of uniform porosity on a substrate; defining a desired pattern on the substrate wherein at least one region of intended lower porosity is defined and wherein at least one region of intended normal porosity is defined; exposing to a flow of a suitable solvent vapor over the region of intended lower porosity wherein the nitrocellulose based film of the region of intended lower porosity is dissolved by the suitable solvent vapor; removing the suitable solvent vapor and the nitrocellulose based film from the region of intended lower porosity wherein the resulted lower porosity region is capable of separating “multiple experiments” that are performed simultaneously over the region of resulted normal porosity.
In one embodiment the suitable solvent vapor is acetone solvent vapor. In one other embodiment exposing the solvent vapor over the region of intended lower porosity is controlled by a robot in a programmable manner. In another embodiment exposing the solvent vapor over the region of intended lower porosity is achieved by delivering the solvent vapor through a flat hypodermic needle.
In one embodiment it further comprises using a shroud to wrap around the needle. In one embodiment it further comprises providing a vacuum to the shroud wherein the vacuum would prevent the solvent vapor from flowing into undesired regions. In one embodiment the vacuum is supplied at a flow rate of 27 normal liters per minute.
In one embodiment the solvent vapor is supplied to the needle generated by bubbling gas through a solvent bottle. In one embodiment the solvent bottle is held at a constant temperature. In one embodiment the bubbling gas is nitrogen air. In one embodiment the nitrogen is bubbled through at pressure of 1.5 psi and at a flow rate of 3.2 normal liters per minute.
In one embodiment the area the region of intended lower porosity can be controlled by altering the speed of the robot. In one embodiment the area the region of intended lower porosity can be controlled by altering the height of the robot over the substrate. In one embodiment the suitable solvent vapor is selected from the group consisting of: acetone, methyl acetate, methyl ether ketone, amyl acetate, chloroform, methylene chloride, ethylacetate, methyl formate, and methyl glycol acetate.
In one embodiment the nitrocellulose film is composed of nitrocellulose and cellulose or polymers. In one embodiment the substrate's temperature is controlled.
In another aspect of the invention, a nitrocellulose film on a substrate with pattern of lower porosity region and high porosity region made by a method comprising: providing a nitrocellulose based film of uniform porosity on a substrate; defining a desired pattern on the substrate wherein at least one region of intended lower porosity is defined and wherein at least one region of intended normal porosity is defined; exposing to a flow of a suitable solvent vapor over the region of intended lower porosity wherein the nitrocellulose based film of the region of intended lower porosity is dissolved by the suitable solvent vapor; removing the suitable solvent vapor and the nitrocellulose based film from the region of intended lower porosity wherein the resulted lower porosity is capable of separating “multiple experiments” that are performed simultaneously over the region of resulted normal porosity.
In one other aspect of the invention, an apparatus for patterning nitrocellulose slide on a substrate wherein the apparatus comprises at least one xyz robot wherein the xyz robot is further comprised of a plurality of pattering heads; at least one solvent bottle for each the pattering head wherein solvent vapor delivery is achieved through bubbling “nitrogen” through the solvent bottle; at least one liquid trap to trap any solvent backflow; at least one nitrocellulose slide on a substrate for patterning; at least one vacuum pump wherein the vacuum pump is used to secure the substrate by suctioning the substrate wherein the vacuum pump is further used to prevent overflow of the solvent vapor into unwanted region; at least one primary electrical solenoid valve to control the solvent vapor; at least one secondary electrical solenoid valve to control the substrate vacuum.
In one embodiment the solvent bottle is maintained at a constant temperature using a warm water bath. In one embodiment it is further comprised of a low-flow regulator used to control gas input pressure at a consistent stable level. In one embodiment it is further comprised of a cold water bath used to cool the substrate to a stable temperature above the dew point.
FIG. 1—A drawing of a patterning head with a needle and a vacuum shroud.
FIG. 2—A drawing a simple system to pattern a nitrocellulose surface using a patterning head mounted on an XYZ robot and solvent delivery by bubbling a gas through a liquid solvent.
FIG. 3—A photograph of a patterned nitrocellulose slide showing patterned regions of low-porosity, transparent nitrocellulose and un-patterned regions of higher porosity, semi-transparent nitrocellulose.
FIG. 4—A photograph of a patterned nitrocellulose slide showing patterned regions of low-porosity, transparent nitrocellulose and un-patterned regions of high porosity, white nitrocellulose.
FIG. 5—A photograph of a patterned nitrocellulose slide showing patterned regions of low-porosity, transparent nitrocellulose and un-patterned circular regions of high porosity, white nitrocellulose.
FIG. 6—Scanning Electron Micrograph (SEM) images showing original un-patterned, high-porosity, white nitrocellulose at 8000× magnification (A) and 1500× magnification (B). Patterned, low porosity, transparent nitrocellulose is shown at 8000× magnification (C) for comparison. The transition region between low-porosity and high-porosity nitrocellulose is shown at 1500× magnification in (D) where the far right of (D) is similar to (B).
FIG. 7—A schematic drawing of a 4 channel patterning head system with independent flow-rate control, temperature stabilized solvent bottles, and temperature controlled substrate platen. Substrates are secured by vacuum to prevent movement.
FIG. 8—Photograph of one embodiment of the 4 channel patterning system.
The present invention relates to the production of defined regions of differing porosity of a nitrocellulose(NC)-based film on a substrate. Starting with a substrate which has a nitrocellulose-based film of uniform porosity, areas of the film are exposed to a flow of a suitable solvent vapor such that the film in the exposed areas are dissolved in the solvent vapor and upon removal of the solvent vapor will dry as a film of lower porosity than the original film. The lower porosity region is then sufficient to separate (usually using a frame with a rubber seal) multiple experiments that are performed simultaneously on the substrate. In order to achieve sharp definition between regions a patterning head as shown in
The invention should not necessarily be limited to only nitrocellulose film but could also apply to mixtures of nitrocellulose and other celluloses or polymers. An example of which might be nitrocellulose mixed with cellulose acetate which is also a common substrate for protein assays.
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This application claims the benefit of priority under 35 U.S.C. 119(e) to the filing date of U.S. provisional patent application No. 61/051,011 entitled “Method for defining regions of differing porosity of a nitrocellulose film on a substrate” which was filed on May 7, 2008, and U.S. provisional patent application No. 61/073,763 entitled “Method for defining regions of differing porosity of a nitrocellulose film on a substrate” which was filed on Jun. 19, 2008 and are incorporated herein by reference.
Number | Date | Country | |
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61051011 | May 2008 | US | |
61073763 | Jun 2008 | US |