Compositions and methods for dehydrated storage of on-board reagents in microfluidic devices
Manufacturing methods and compositions are described for production of self-contained microfluidic cartridge devices with on-board reagents for molecular biological testing. Sensitive reagents are stored in dry form without lyophilization or freezing, and reconstituted at the point of use with either a biological sample or a sample eluate at the point of use. Manufacturing methods include sheet and roll fabrication processes where the reagents are printed in place and sealed within individual microfluidic cartridges before gel vitrification.
1. A method for stabilizing a DNA polymerase for storage in a gel-like glass above the freezing temperature of water without lyophilization in a chamber or channel of a microfluidic device, the method comprising:
a) combining said DNA polymerase with an aqueous solution composed of:
i) from about 1.0% w/v to 10% w/v of a disaccharide or a trisaccharide;
ii) optionally from about 0.001% w/v to 0.1% w/v of a high molecular weight polyethylene glycol;
iii) from about 0.001% w/v to 0.3% w/v of a fluorosurfactant;
iv) optionally from about 0.001% w/v to about 1.0% w/v of an amino acid;
iv) optionally from about 0.1 mg/mL to 10 mg/mL of a carrier protein; and
v) optionally a compatible buffer, thereby forming a composite glass precursor solution;
b) depositing in said microfluidic device a droplet of said composite glass precursor solution containing a quantity of said DNA polymerase effective for said assay;
c) drying said droplet at controlled room temperature to form a gel spot on said surface; and
d) then closing and sealing said gel spot in said microfluidic cartridge in a gas tight pouch under a dry atmosphere with a dessicant, said dessicant further vitrifying said gel spot during storage.
2. The method of claim 1 , wherein said disaccharide is selected from trehalose and said trisaccharide is selected from melezitose or raffinose.
3. The method of claim 1 , wherein said high molecular weight polyethylene glycol has a molecular weight of 0.1 to 5 MDa and is a linear chain polyoxyethylene glycol or a branched chain polyoxyethylene glycol.
4. The method of claim 1 , wherein said carrier protein is bovine serum albumin or fish gelatin.
5. The method of claim 1 , wherein said fluorosurfactant is a non-ionic fluoroalkylsurfactant.
6. The method of claim 1 , wherein said fluorosurfactant is Fluorosurfactant FC-4430.
7. The method of claim 1 , further comprising a step for passivating said chamber or channel prior to depositing said droplet.
8. The method of claim 1 , wherein controlled room temperature is about 20° C.