Preparation of glassified biological reagents
A biological, reagent technology, applied in the direction of fixation on/in organic carriers, powder conveying, etc., can solve problems such as trouble, manual labor, etc.
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Embodiment 1
[0086] Example 1: Preparation of Dried PCR Mixtures in 384-well Polystyrene Plates
[0087] Biological reagent preparations, in this case reagent mixes for PCR, were prepared following standard protocols. Briefly, a formulation of 10 μl of spheres contained 25 mM Tris-HCl (pH 9.0 at room temperature), 125 mM KCl, 3.75 mM MgCl 2 , 0.5mM dNTP, 0.6mg / ml BSA, 3.5 units of rTaq DNA polymerase and synthetic polymers including Ficoll 400 (6.25%), Ficoll 70 (6.25%) and the second carbohydrate melezitose (10%) glass-forming filler material. All reagents were generally autoclaved or filter sterilized prior to use. Reagents were prepared and stored on ice until mixed and dispensed into polystyrene plates or silica molds. The final formulation consisted of the glass-forming filler material, BSA, dNTPs and rTaq DNA polymerase, and the above salts.
[0088] The reagent homogeneous solution has a final volume of 10 μl per dose. This allows for a PCR working volume of 25 μl. Use an auto...
Embodiment 2
[0093] Example 2: Preparation of Dried PCR Mixtures in 96 Well Silica Molds
[0094] Biological reagent formulations and glass-forming fill materials were prepared as in Example 1 and mixed together. Use an automatic pipette to dispense approximately 20 μl of the reagent mixture into a 96-well silica mold ( image 3, upper left). Reagents were frozen by placing the silica molds in a pre-cooled (-46°C) Vertis freeze dryer for approximately 60 minutes. The frozen reagents were then subjected to a first and then a second drying process as described in Table 2 above.
[0095] Dried reagents can be stored in the mold as a complete package by simply covering with an adhesive seal and by placing the plate in an aluminum pouch containing the desiccant. Alternatively, the dried reagents are removed from the mold as a dried reagent cake, which is then stored in a bottle containing a desiccant.
[0096] The stability of the dried reagent composition was tested by real-time PCR amplif...
Embodiment 3
[0098] Example 3: Preparation of Dried PCR Mixtures in 96-well Polystyrene Plates
[0099] Biological reagent formulations and glass-forming fill materials were prepared as in Example 1 and mixed together. Dispense 10 μl of reagent mix using a liquid dispensing robot ( figure 1 , lower left). Place the 96-well plate on a 96-well metal stand ( Figure 4 ). Reagents were frozen by placing the 96-well plate with the metal rack in a pre-cooled (-46°C) Vertis freeze dryer for approximately 60 minutes. The frozen reagents were then subjected to a first and then a second drying process as described in Table 2 above.
[0100] Dried reagents can be stored in the 96-well plate as a complete package by simply covering the plate with an adhesive seal or cap or heat seal with the aid of a heat sealer. Lambda DNA real-time PCR functional tests were performed using freshly prepared "wet" preparations and dried reagents together with puRe Taq RTG beads. Sealed plates were stored in bag...
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