PREFILLED CARTRIDGE

Abstract

The disclosure pertains to a microfluidic cartridge comprising at least one microchannel and at least a set of functionalized microcarriers, the microcarriers being localized within the microchannel, wherein the functionalized microcarriers are coated with at least a lyoprotectant. The disclosure further pertains to a process of manufacture of a microfluidic cartridge according to the invention, said process comprising: providing a microfluidic cartridge comprising at least one microchannel and at least a set of functionalized microcarriers, preferably in suspension in a buffer solution, the microcarriers being localized within the microchannel; flowing a stabilizing buffer into the at least one microchannel and incubating the functionalized microcarriers with said stabilizing buffer for at least 10 minutes, wherein the stabilizing buffer is a composition comprising a lyoprotectant, preferably wherein the lyoprotectant is chosen from the list consisting of sugars and sugar alcohols and mixtures thereof; and drying the at least one microchannel.

Claims

1-19. (canceled)

20. A process of manufacture of a microfluidic cartridge, the micro fluidic cartridge comprising: at least one microchannel, wherein the microchannel is a hollow structure configured for the passage of fluids; and at least a set of functionalized microcarriers, wherein the microcarriers are localized within the microchannel, and wherein the functionalized microcarriers are coated with at least a lyoprotectant; said process comprising: providing a microfluidic cartridge comprising at least one microchannel and at least a set of functionalized microcarriers in suspension in a buffer solution, the microcarriers being localized within the microchannel; flowing a stabilizing buffer into the at least one microchannel and incubating the functionalized microcarriers with said stabilizing buffer for at least 10 minutes, wherein the stabilizing buffer is a composition comprising the lyoprotectant; and drying the at least one microchannel.

21. The process of claim 20, wherein the lyoprotectant is or comprises a sugar selected from the list consisting of sucrose, trehalose, sorbose, stachyose, gentianose, melezitose, raffinose, fructose, apiose, mannose, maltose, isomaltulose, lactose, lactulose, arabinose, xylose, lyxose, digitoxose, fucose, quercitol, allose, altrose, primeverose, ribose, rhamnose, galactose, glyceraldehyde, tagatose, turanose, sophorose, maltotriose, manninotriose, rutinose, scillabiose, cellobiose, gentiobiose, glucose, cellulose and cellulose derivatives, hydroxyethylstarch, soluble starches, dextrans, highly branched, high-mass, hydrophilic polysaccharides.

22. The process of claim 20, wherein the lyoprotectant is or comprises a sugar-alcohol selected from the list consisting of lactitol, mannitol, maltitol, xylitol, erythritol, myoinositol, threitol, sorbitol, and glycerol.

23. The process of claim 20, wherein the stabilizing buffer is flown in the at least one microchannel at room temperature for more than 30 seconds.

24. The process of claim 20, wherein the microcarriers are incubated in the presence of said stabilizing buffer at room temperature for at least 10 minutes.

25. The process of claim 20, wherein the step of drying the at least one microchannel comprises a step of removing part of the stabilizing buffer from the at least one microchannel by flushing said microchannel with a gas under pressure.

26. The process of claim 20, wherein the step of drying the at least one microchannel comprises a step of removing part of the stabilizing buffer from the at least one microchannel by vacuum drying.

27. The process of claim 20, wherein the step of drying the at least one microchannel comprises absorbing the stabilizing buffer.

28. The process of claim 25, wherein the step of removing part of the stabilizing buffer from the at least one microchannel by flushing said microchannel with a gas under pressure is followed by a step of incubating the microfluidic cartridge in a closed chamber, in the presence of dry air.

29. The process of claim 25, wherein the step of drying the at least one microchannel comprises removing the stabilizing buffer by flushing said microchannel with a gas under a positive differential pressure of at least 20 mBar.

30. The process of claim 26, wherein the step of drying the at least one microchannel comprises a step of vacuum drying at an absolute pressure between 40 mBar and 700 mBar.

31. The process of claim 30, wherein the at least one microchannel is dried until the humidity rate of the air inside the vacuum drying equipment reaches between 0.5% and 20%.

32. The process of claim 27, wherein absorbing the stabilizing buffer comprises use of an absorbing material positioned at one extremity of the microchannel.

33. The process of claim 20, further comprising a step of packing the microfluidic cartridge in a container.

34. The process of claim 33, wherein the container is vacuum sealed after the microfluidic cartridge is packed in the container.

35. The process of claim 33, wherein the container comprises a desiccant.

36. The process of claim 20, wherein the surface of the microcarriers and the internal surface of the microchannel are coated with said lyoprotectant.

37. The process of claim 20, wherein the at least one set of microcarriers are functionalized with a detection molecule, wherein said detection molecule is selected from the list consisting of a protein, a peptide, a DNA fragment, a RNA fragment and a ssDNA fragment.

38. The process of claim 20, wherein the microfluidic cartridge comprises more than one microchannel, and wherein each of the microchannels of the microfluidic cartridge comprises at least 2, 3, 4, 5, 6, 10, 20 sets of microcarriers.

39. A microfluidic cartridge produced by the process according to claim 20.

Description

FIGURE LEGEND

[0100] FIG. 1: Stability of pre-filed cartridge after storage at different temperatures.

[0101] Prefilled cartridges comprising microcarriers functionalized with either an anti-IL-4 antibody (set 1), an anti-II-6 antibody (set 2), an anti-IL-8 antibody (set 3), an anti-TNF-alpha antibody (set 4), an alternative anti-TNF-alpha antibody, different from the set 4 antibody (set 5) or unfunctionalized (set 6) were prepared. They were then stored at 20 C., 4 C., 25 C. or 37 C., and their functionality was tested at different storage time. The functional test consisted in measuring fluorescence in calibrated conditions, and comparing the fluorescence obtained with that obtained with a cartridge of reference.

EXAMPLES

[0102] Prefilled cartridges were prepared as follows:

[0103] Empty cartridges comprising microchannels were provided, as well as conventional unfunctionalized microcarriers. Conventional unfunctionalized microcarriers typically harbor chemical moieties (such as streptavidine molecules or carboxyl functions) on their surface so as to enable functionalization with a molecule of interest. Unfunctionalized microcarriers were introduced in the microchannels using conventional techniques. The functionalization of the microcarriers was then performed directly within the microchannels.

[0104] Several types of functionalization were tested in separate cartridges. Accordingly: [0105] In a first set of cartridge, the microchannels were flushed with a buffer comprising 1 g/mL of anti-IL-4 antibody (set 1), [0106] In a second set of cartridge, the microchannels were flushed with a buffer comprising 1 g/mL of anti-IL-6 antibody (set 2), [0107] In a third set of cartridge, the microchannels were flushed with a buffer comprising 1 g/mL of anti-IL-8 antibody (set 3), [0108] In a fourth set of cartridge, the microchannels were flushed with a buffer comprising 1 g/mL of anti-TNF-alpha antibody (set 4), [0109] In a fifth set of cartridge, the microchannels were flushed with a buffer comprising 1 g/mL of an alternative anti-TNF-alpha antibody, different from the set 4 antibody (set 5).

[0110] A sixth set of microcarriers was left unfunctionalized (set 6).

[0111] Functionalization of the microcarriers was performed by incubating the microchannels during about 30 minutes to 60 minutes. After incubation, the microchannels were flushed with PBST buffer for one minute, to rinse the antibody. The microchannels were then flushed with stabilizing buffer (Coating Stabilizer and Blocking Buffer commercialized by the company Meridian Life Science). The microchannels and thus the microcarriers were left to incubate in presence of stabilizing buffer for one hour at room temperature.

[0112] The microchannels were then dried by: [0113] gas purge, using compressed air at 0.4 bar for 3 minutes., [0114] followed by evaporation under vacuum for 15 hours at room temperature in a vacuum chamber at around 0.71 bar.

[0115] The cartridges were then packed in aluminum bags with silicagel (2 g), under 20% vacuum.

[0116] The cartridges were stores at 4 C., 20 C., 25 C. or 37 C., and their functionality was tested at various storage time. The functional test consisted in measuring fluorescence in calibrated conditions, and comparing the fluorescence obtained with that obtained with a cartridge of reference. In each case, the cartridge of reference was a readily made cartridge with microcarriers functionalized with the same antibody as used in the cartridge to be tested. The ratios of fluorescence obtained in different conditions are presented in FIG. 1. The cartridge of the invention can be stored for extended periods of time at any of the temperature tested without any significant loss of functionalization of the microcarriers.