Optimized surfactant formulations for droplet-based microfluidics applications

Abstract

The present invention is directed to formulations comprising a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant of formula A-B and a triblock co-polymer surfactant of formula B'-A'-B', wherein A and A' independently of each other represent a water-soluble polymer block having a weight average molecular weight in the range of 300 g / mol to 2000 g / mol, and B and B' represent independently of each other a perfluorinated polyether block having a weight average molecular weight in the range of 1500 g / mol to 7500 g / mol. Said formulations exhibit improved properties which render them particularly useful in droplet-based microfluidics applications, including the construction of lipid vesicles, cell encapsulation and monitoring, and droplet digital molecular biology.

Description

[0001] Optimized surfactant formulations for droplet-based microfluidics applications

[0002] Specification

[0003] The present invention is related to formulations comprising a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant of formula A-B and a triblock co-polymer surfactant of formula B’-A’-B’, wherein A and A’ independently of each other represent a water-soluble polymer block having a weight average molecular weight in the range of 300 g / mol to 2000 g / mol, and B and B’ represent independently of each other a perfluorinated polyether block having a weight average molecular weight in the range of 1500 g / mol to 7500 g / mol. Said formulations exhibit improved properties which render them particularly useful in droplet-based microfluidics applications, including the construction of lipid vesicles, cell encapsulation and monitoring, and droplet digital molecular biology.

[0004] Background of the invention

[0005] Microfluidic technology facilitates the precise manipulation and control of fluid flows ranging from microliters to femtoliters within channels of micrometer dimensions. This technology leverages the distinct behavior of fluids at the microscale, which arises from variations in physical properties compared to their behavior at larger scales. Such capabilities make microfluidics an invaluable tool across multiple disciplines, including engineering, physics, chemistry, biochemistry, nanotechnology, synthetic biology, and biotechnology.

[0006] In the domain of droplet-based microfluidics, this technology employs immiscible fluids to generate an emulsion, thereby forming discrete reaction vessels, called droplets. The stabilization of water-in-oil (w / o) droplets is addressed through the mitigation of the inherent high Laplace pressure that prompts immediate fusion of droplets postproduction, aiming to minimize surface free energy. This stabilization is achieved by diminishing the interfacial tension between the water and fluorinated oil phases. The method involves the application of surfactants, which are amphiphilic molecules that naturally congregate at the interface of two immiscible fluids, thereby effectively reducing the interfacial tension. This approach ensures the maintenance of droplet integrity and prevents coalescence, contributing to the stability of the emulsion system.

[0007] Droplet-based microfluidics can be used for a wide range of types of experiments in biotechnology, for example in the construction of lipid vesicles, to encapsulate and monitor cells and for droplet digital molecular biology. Already these three types of

[0008] GAR-P04610W010 Application (final). docx experiments have different requirements on the physicochemical and biochemical properties of the surfactants. In lipid vesicle construction, transitioning lipid vesicles from an oil / water to a water / water system requires a surfactant that can create a metastable system, robust enough for stabilization of w / o droplets but also capable of being destabilized for vesicle release. For cell encapsulation, additional nutrients must be included for the survival and functionality of the encapsulated cells. This creates a high osmotic pressure, which increases the fusion forces of the droplets. In droplet digital molecular biology, droplets have to maintain stability during chemical reactions and analyses, to ensure accurate results. This includes significant shear stress from physical manipulations and high reaction temperatures.

[0009] Accordingly, different fluorosurfactants or fluorosurfactant formulations for specific experiment of droplet-based microfluidics conditions are needed to ensure the quality of the respective experiments.

[0010] International patent application WO 2014 / 138711 A1 discloses surfactant compositions and emulsions designed for use in droplet microfluidic systems. The disclosed surfactants having a PFPE-PEG-PFPE triblock polymer structure , where the PFPE blocks are based on Krytox® 157 FSH linked by amide groups, enable the formation of stable water-in-oil emulsions with enhanced droplet integrity and biocompatibility, suitable for biological and diagnostic applications in microfluidic environments.

[0011] The article Marian Weiss et al. (Nature Materials 2017, 17(1 ), p. 89-96) discloses a method for the sequential bottom-up assembly of synthetic cell-like compartments using microfluidics, particularly the creation of droplet-stabilized giant unilamellar vesicles (dsGUVs), which offer improved mechanical and chemical stability compared to conventional lipid-based protocells. These compartments are formed by encapsulating lipid vesicles within copolymer-stabilized water-in-oil droplets, allowing precise manipulation and loading of biomolecules via pico-injection technology The article further reports on the production of three fluorosurfactant denominated as PFPE(7000 g / mol)-PEG(1400 g / mol)-PFPE(7000 g / mol), PFPE(2500 g / mol)-PEG(600 g / mol)- PFPE(2500 g / mol) and Au-PEG(436 g / mol)-PFPE(7000 g / mol). 3 pM of the gold linked diblock surfactant and 2.5 mM of the triblock surfactants are mixed with FC-40.

[0012] Holtze C. et al. (Lab on a Chip 2008, 8(10), p. 1632) report on the synthesis and application of biocompatible fluorosurfactants designed to stabilize water-in- fluorocarbon emulsions used in microfluidic systems, particularly for biological assays. The surfactants are synthesized by coupling perfluoropolyether (PFPE) with polyethylene glycol (PEG) via amide bonds, forming diblock (PFPE-PEG) and triblock (PFPE-PEG-PFPE) copolymers. The PFPE segments impart fluorophilicity for

[0013] GAR-P04610W010 Application (final). docx solubility in fluorocarbon oils, while the PEG segments prevent biomolecular adsorption, ensuring compatibility with encapsulated biological components.

[0014] Scanga Randall et al. (RSC Advances 2018, 8(23) p. 12960-12974) report on the design and synthesis of PEG-based fluorinated surfactants for use in droplet microfluidics. The study focuses on improving the accessibility and structural diversity of PFPE-PEG surfactants through click chemistry approaches, specifically copper- catalyzed azide-alkyne cycloaddition (CuAAC) and thiol-yne reactions. The invention overcomes limitations in traditional PFPE-PEG synthesis, namely its complexity and moisture sensitivity, by introducing a stable, click-ready PFPE propargyl intermediate that can be efficiently functionalized with various hydrophilic heads (e.g., PEG and glycerol ethoxylates).

[0015] Thus, it is the objective of the present invention to provide improved fluorosurfactant formulations for droplet-based microfluidics applications.

[0016] The objective of the present invention is solved by the teachings of the independent claims. Further advantageous features, aspects and details of the invention are evident from the dependent claims, the description, the figures, and the examples of the present application.

[0017] Brief description of the invention

[0018] The present invention provides tailor-made surfactant formulations comprising a water- immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant of formula A-B and a triblock co-polymer surfactant of formula B’-A’-B’, wherein A and A’ independently of each other represent a water-soluble polymer block having a range in molecular weight from 300 g / mol to 2000 g / mol, and B and B’ represent independently of each other a perfluorinated polyether block having a range in molecular weight from 1500 g / mol to 7500 g / mol, wherein the weight average molecular weights of A, A’, B, and B’ are determined by gel permeation chromatography, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9 have the following formula: surfactant 1 : PFPE(2000 g / mol-L1-PEG(350 g / mol) surfactant 2: PFPE(4000 g / mol)-L1-PEG(350 g / mol) surfactant 3: PFPE(7000 g / mol)-L1-PEG(350 g / mol) surfactant 4: PFPE(2000 g / mol)-L1-PEG(550 g / mol)

[0019] GAR-P04610W010 Application (final). docx ABK

[0020] 4 surfactant 5: PFPE(4000 g / mol)-L1-PEG(550 g / mol) surfactant 6: PFPE(7000 g / mol)-L1-PEG(550 g / mol) surfactant 7: PFPE(2000 g / mol)-L1-PEG(750 g / mol) surfactant 8: PFPE(4000 g / mol)-L1-PEG(750 g / mol) surfactant 9: PFPE(7000 g / mol)-L1-PEG(750 g / mol) wherein L1represents a triazole ring having the structure: and wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein surfactant 10 to surfactant 18 have the following formula: surfactant 10: PFPE(2000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(2000 g / mol) surfactant 11 : PFPE(4000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(4000 g / mol) surfactant 12: PFPE(7000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(7000 g / mol) surfactant 13: PFPE(2000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(2000 g / mol) surfactant 14: PFPE(4000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(4000 g / mol) surfactant 15: PFPE(7000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(7000 g / mol) surfactant 16: PFPE(2000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(2000 g / mol) surfactant 17: PFPE(4000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(4000 g / mol) surfactant 18: PFPE(7000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(7000 g / mol) wherein L2and L3represent a triazole ring having the structures:

[0021] In one embodiment of the present invention, these formulations include different ratios of the at least two amphiphilic polymers independently selected from a diblock copolymer surfactant of formula A-B and a triblock co-polymer surfactant of formula B’-A’-B’.

[0022] The diblock co-polymer surfactant has a general structure of A-B, wherein A is a water- soluble polymer like polyethylene glycol and B is a perfluorinated polymer like

[0023] GAR-P04610W010 Application (final). docx perfluoropolyether. The polymer blocks A and B may be connected by an ether group, an ester group, an amine group, an amide group or a triazole ring. According to the present invention, the polymer A has a range in molecular weight from 300 g / mol to 2000 g / mol and the polymer B has a range in molecular weight from 1500 g / mol to 7500 g / mol.

[0024] The triblock co-polymer surfactant has a general structure of B’-A’-B’, wherein A’ is a water-soluble polymer like polyethylene glycol and B’ is a perfluoropolyether. The polymer blocks B’ and A’ may be connected by an ether group, an ester group, an amine group, an amide group or a triazole ring. According to the present invention, the polymer A’ has a range in molecular weight from 300 g / mol to 2000 g / mol and the polymer B’ has a range in molecular weight from 1500 g / mol to 7500 g / mol.

[0025] The efficiency of how much the surfactants is stabilizing the w / o droplets was determined by the measuring the reduction of interfacial tension using pendent drop methodology and diameter distribution of w / o droplets over time.

[0026] In a preferred embodiment of the inventive formulation, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7000 (1 , 1 ,1 , 2, 2,3,3- heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 ,1 , 1 ,2,3, 3-hexafluoro- 3-methoxy-2-(trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1-ethoxy- 1 , 1 ,2,2,3,3,4,4,4-nonafluorobutane and 1 -ethoxy-1 , 1 , 2,3,3, 3-hexafluoro-2-

[0027] (trifluoromethyl)propane), HFE-7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3-methoxy-4-

[0028] (trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2- (trifluoromethyl)hexane), FC-40 (1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N,N- bis(nonafluorobutyl)butan-1 -amine), FC-43 (1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N, N- bis(1 ,1 ,2,2,3,3,4,4,4-nonafluorobutyl)butan-1-amine), FC-70 (1 ,1 , 2, 2, 3, 3, 4, 4, 5,5,5- undecafluoro-N,N-bis(undecafluoropentyl)pentan-1 -amine), FC-72

[0029] (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned.

[0030] In a preferred embodiment of the inventive formulation, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 ,1 , 1 ,2,3, 3-hexafluoro-3- methoxy-2-(trifluoromethyl)propane), HFE-7500 (3-ethoxy-1 ,1 ,1 , 2, 3, 4, 4, 5, 5, 6,6,6- dodecafluoro-2-(trifluoromethyl)hexane), and FC-40 (1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N, N- bis(nonafluorobutyl)butan-1 -amine).

[0031] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock copolymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the total concentration of the least one amphiphilic polymer is in a range between 0.5 mM x a + 0.5 mM and 10 mM x a + 10 mM, wherein a is the molar fraction of the diblock co-polymer surfactant in the least one amphiphilic polymer.

[0032] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock copolymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the total concentration of the least one amphiphilic polymer is 2.5 mM x a + 2.5 mM, wherein a is the molar fraction of the diblock co-polymer surfactant in the least one amphiphilic polymer.

[0033] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and two amphiphilic polymers selected from a diblock co-polymer surfactant, and a triblock co-polymer surfactant, wherein the diblock-co-polymer is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9 and the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein surfactant 10 to surfactant 18, wherein surfactant 1 to surfactant 18 have the meanings as defined herein.

[0034] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and three amphiphilic polymers selected from two different diblock co-polymer surfactants and a triblock co-polymer surfactant or one diblock co-polymer surfactant and two different triblock co-polymer surfactants wherein the diblock-co-

[0035] GAR-P04610W010 Application (final). docx polymer is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9 and the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein surfactant 10 to surfactant 18, wherein surfactant 1 to surfactant 18 have the meanings as defined herein.

[0036] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and four amphiphilic polymers selected from three different diblock co-polymer surfactants and one triblock co-polymer surfactant, or four different diblock co-polymer surfactants, or two different diblock co-polymer surfactants and two different triblock co-polymer surfactants, or one diblock co-polymer surfactant and three different triblock co-polymer surfactants, or four different triblock co-polymer surfactants, wherein the diblock-co-polymer is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9 and the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein surfactant 10 to surfactant 18, wherein surfactant 1 to surfactant 18 have the meanings as defined herein.

[0037] According to the present invention, these formulations are not contaminated with small amounts of free perfluoropolyether carboxylic acid. Commercially available diblock copolymer surfactants or triblock co-polymer surfactants comprising a perfluorinated polyether block commonly contain small amounts of perfluoropolyether carboxylic acids as by-products of their synthetic preparation. As the perfluoropolyether carboxylic acids have a significant influence on the droplet formation and droplet properties, including droplet stability, impurities of perfluoropolyether carboxylic acids, such as Krytox™ in the amphiphilic polymer effect the droplet formation and stabilization in an unforeseeable manner. Therefore, commercially available diblock co-polymer surfactants or triblock co-polymer surfactants comprising a perfluorinated polyether block are not suitable amphilic polymers for the tailor-made surfactant formulations of the present invention for droplet-based microfluidics applications, including the construction of lipid vesicles, cell encapsulation and monitoring, and droplet digital molecular biology.

[0038] Thus, the present invention is also directed to a formulation comprising a water- immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant, wherein the diblock co-polymer surfactant is selected from the

[0039] GAR-P04610W010 Application (final). docx group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, and wherein the formulation is devoid of free perfluoropolyether carboxylic acid.

[0040] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant s, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the formulation is devoid of free perfluoropolyether carboxylic acid, and wherein the perfluoropolyether carboxylic acid is a polymer of hexafluoropropylene oxide.

[0041] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent, at least one amphiphilic polymer, and 0.1 mM to 15 mM of a perfluoropolyether carboxylic acid, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18.

[0042] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent, at least one amphiphilic polymer, and 0.1 mM to 15 mM of a perfluoropolyether carboxylic acid, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15,

[0043] GAR-P04610W010 Application (final). docx surfactant 16, surfactant 17, and surfactant 18, and wherein the perfluoropolyether carboxylic acid is a polymer of hexafluoropropylene oxide.

[0044] The present invention further relates tailor-made surfactant formulations for the construction of lipid vesicles. According to a more specific embodiment, the surfactant formulation is optimized in regards of lipid vesicle release yield. According to a preferred embodiment, the surfactant formulation comprises a mixture of a diblock co-polymer surfactant and a triblock co-polymer surfactant. The triblock co-polymer surfactant could be in a molecular ratio of 37.5 %. The diblock co-polymer surfactant could be in a molecular ratio of 62.5 %. In further preferred embodiments, the formulation may also contain more than two amphiphilic polymers.

[0045] The present invention further relates to a tailor-made surfactant formulation for encapsulating and monitoring cells. According to a more specific embodiment, the surfactant formulation is optimized in regards of cell viability over multiple days. According to a preferred embodiment, the surfactant formulation comprises a mixture of a diblock co-polymer surfactant and a triblock co-polymer surfactant. The triblock copolymer surfactant could be in a molecular ratio of 37.5 %. The diblock co-polymer surfactant could be in a molecular ratio of 62.5 %. In further preferred embodiments, the formulation may also contain more than two amphiphilic polymers.

[0046] The present invention further relates to a tailor-made surfactant formulation for droplet digital molecular biology. According to a more specific embodiment, the surfactant formulation is optimized in regards of fluorescent and colorimetric readout. According to a preferred embodiment, the surfactant formulation comprises a mixture of a diblock copolymer surfactant and a triblock co-polymer surfactant. The triblock co-polymer surfactant could be in a molecular ratio of 37.5%. The diblock co-polymer surfactant could be in a molecular ratio of 62.5%. In further preferred embodiments, the formulation may also contain more than two amphiphilic polymers.

[0047] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer wherein formulation comprises the following diblock co-polymer surfactants and triblock co-polymer surfactants selected from surfactant 1 surfactant 1 , surfactant 10, surfactant 11 surfactant 1 , surfactant 10, surfactant 17 surfactant 1 , surfactant 11 , surfactant 13, surfactant 15 surfactant 1 , surfactant 13, surfactant 15, surfactant 16 surfactant 1 , surfactant 2, surfactant 4, surfactant 15 surfactant 1 , surfactant 3, surfactant 7, surfactant 11 surfactant 1 , surfactant 4, surfactant 11 , surfactant 13

[0048] GAR-P04610W010 Application (final). docx surfactant 1 , surfactant 4, surfactant 11 , surfactant 16 surfactant 1 , surfactant 4, surfactant 18 surfactant 1 , surfactant 6, surfactant 11 , surfactant 12 surfactant 1 , surfactant 7, surfactant 11 , surfactant 12 surfactant 1 , surfactant 7, surfactant 13, surfactant 14 surfactant 1 , surfactant 8, surfactant 15, surfactant 18 surfactant 1 , surfactant 9, surfactant 13, surfactant 16 surfactant 10 surfactant 10, surfactant 13, surfactant 14, surfactant 15 surfactant 10, surfactant 14, surfactant 17, surfactant 18 surfactant 11 surfactant 11 , surfactant 6, surfactant 12 surfactant 11 , surfactant 9, surfactant 16 surfactant 12 surfactant 12, surfactant 13, surfactant 17 surfactant 13 surfactant 13, surfactant 14, surfactant 18 surfactant 13, surfactant 15 surfactant 13, surfactant 7, surfactant 9 surfactant 14 surfactant 14, surfactant 10, surfactant 17 surfactant 14, surfactant 3, surfactant 15 surfactant 15 surfactant 15, surfactant 12, surfactant 14 surfactant 15, surfactant 14, surfactant 18 surfactant 16 surfactant 16, surfactant 14 surfactant 16, surfactant 8, surfactant 18 surfactant 17 surfactant 17, surfactant 5, surfactant 13 surfactant 17, surfactant 8, surfactant 18 surfactant 17, surfactant 8, surfactant 9 surfactant 18 surfactant 18, surfactant 1 , surfactant 13 surfactant 18, surfactant 10, surfactant 13 surfactant 18, surfactant 14, surfactant 16 surfactant 18, surfactant 5, surfactant 11 surfactant 18, surfactant 5, surfactant 12 surfactant 18, surfactant 9, surfactant 13 surfactant 2, surfactant 11 , surfactant 18 surfactant 2, surfactant 3, surfactant 4, surfactant 10 surfactant 2, surfactant 3, surfactant 4, surfactant 12 surfactant 2, surfactant 3, surfactant 8 surfactant 2, surfactant 4, surfactant 5, surfactant 12 surfactant 2, surfactant 4, surfactant 8, surfactant 12 surfactant 2, surfactant 5, surfactant 18 surfactant 2, surfactant 6, surfactant 15, surfactant 16 surfactant 3, surfactant 1 , surfactant 15 surfactant 3, surfactant 10, surfactant 11 surfactant 3, surfactant 10, surfactant 12, surfactant 16 surfactant 3, surfactant 15, surfactant 16 surfactant 3, surfactant 2, surfactant 7

[0049] GAR-P04610W010 Application (final). docx surfactant 3, surfactant 4, surfactant 10, surfactant 13 surfactant 3, surfactant 4, surfactant 16 surfactant 3, surfactant 4, surfactant 5, surfactant 16 surfactant 3, surfactant 4, surfactant 8 surfactant 3, surfactant 7, surfactant 15 surfactant 3, surfactant 8, surfactant 14, surfactant 16 surfactant 4, surfactant 5, surfactant 6 surfactant 5, surfactant 12, surfactant 16 surfactant 5, surfactant 13, surfactant 17 surfactant 5, surfactant 14 surfactant 5, surfactant 6, surfactant 11 , surfactant 15 surfactant 5, surfactant 7, surfactant 12, surfactant 14 surfactant 6 surfactant 6, surfactant 13, surfactant 17 surfactant 6, surfactant 15 surfactant 7 surfactant 7, surfactant 1 surfactant 7, surfactant 1 , surfactant 10 surfactant 7, surfactant 10, surfactant 17 surfactant 7, surfactant 11 , surfactant 12, surfactant 16 surfactant 7, surfactant 12, surfactant 13, surfactant 17 surfactant 7, surfactant 13, surfactant 14, surfactant 17 surfactant 7, surfactant 16 surfactant 7, surfactant 8, surfactant 13, surfactant 15 surfactant 8 surfactant 8, surfactant 1 , surfactant 18 surfactant 8, surfactant 10, surfactant 11 , surfactant 12 surfactant 8, surfactant 11 , surfactant 14, surfactant 15 surfactant 8, surfactant 12, surfactant 14, surfactant 15 surfactant 8, surfactant 17 surfactant 8, surfactant 3, surfactant 15 surfactant 8, surfactant 9, surfactant 10, surfactant 14 surfactant 9 surfactant 9, surfactant 11 , surfactant 12, surfactant 16 surfactant 9, surfactant 17 surfactant 9, surfactant 18 surfactant 9, surfactant 2, surfactant 4.

[0050] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer wherein formulation comprises the following diblock co-polymer surfactants and triblock co-polymer surfactants selected from surfactants molar ratio surfactant 1 surfactant 1 , surfactant 10, surfactant 11 2:1 :1 surfactant 1 , surfactant 10, surfactant 17 2:1 :1 surfactant 1 , surfactant 2, surfactant 4, surfactant 15 1 : 1 : 1 : 1 surfactant 1 , surfactant 3, surfactant 7, surfactant 11 1 : 1 : 1 : 1 surfactant 1 , surfactant 4, surfactant 11 , surfactant 13 1 : 1 : 1 : 1

[0051] GAR-P04610W010 Application (final). docx ABK

[0052] 12 surfactant 1 , surfactant 4, surfactant 11 , surfactant 16 1 : 1 : 1 : 1 surfactant 1 , surfactant 4, surfactant 18 2:1 :1 surfactant 1 , surfactant 6, surfactant 11 , surfactant 12 1 : 1 : 1 : 1 surfactant 1 , surfactant 7, surfactant 11 , surfactant 12 1 : 1 : 1 : 1 surfactant 1 , surfactant 7, surfactant 13, surfactant 14 1 : 1 : 1 : 1 surfactant 1 , surfactant 8, surfactant 15, surfactant 18 1 : 1 : 1 : 1 surfactant 1 , surfactant 9, surfactant 13, surfactant 16 1 : 1 : 1 : 1 surfactant 11 , surfactant 6, surfactant 12 2:1 :1 surfactant 13 surfactant 13, surfactant 7, surfactant 9 2:1 :1 surfactant 14, surfactant 3, surfactant 15 2:1 :1 surfactant 17, surfactant 5, surfactant 13 2:1 :1 surfactant 17, surfactant 8, surfactant 9 2:1 :1 surfactant 18, surfactant 5, surfactant 11 2:1 :1 surfactant 18, surfactant 5, surfactant 12 2:1 :1 surfactant 2, surfactant 11 , surfactant 18 2:1 :1 surfactant 2, surfactant 3, surfactant 4, surfactant 10 1 : 1 : 1 : 1 surfactant 2, surfactant 3, surfactant 4, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 3, surfactant 8 2:1 :1 surfactant 2, surfactant 4, surfactant 5, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 4, surfactant 8, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 5, surfactant 18 2:1 :1 surfactant 2, surfactant 6, surfactant 15, surfactant 16 1 : 1 : 1 : 1 surfactant 3, surfactant 1 , surfactant 15 2:1 :1 surfactant 3, surfactant 4, surfactant 10, surfactant 13 1 : 1 : 1 : 1 surfactant 3, surfactant 4, surfactant 5, surfactant 16 1 : 1 : 1 : 1 surfactant 3, surfactant 4, surfactant 8 2:1 :1 surfactant 3, surfactant 7, surfactant 15 2:1 :1 surfactant 3, surfactant 8, surfactant 14, surfactant 16 1 : 1 : 1 : 1 surfactant 4, surfactant 5, surfactant 6 2:1 :1 surfactant 5, surfactant 12, surfactant 16 2:1 :1 surfactant 5, surfactant 13, surfactant 17 2:1 :1 surfactant 5, surfactant 14 1 :1 surfactant 5, surfactant 6, surfactant 11 , surfactant 15 1 : 1 : 1 : 1 surfactant 5, surfactant 7, surfactant 12, surfactant 14 1 : 1 : 1 : 1 surfactant 6 surfactant 6, surfactant 13, surfactant 17 2:1 :1 surfactant 6, surfactant 15 1 :1 surfactant 7 surfactant 7, surfactant 1 3:1 surfactant 7, surfactant 1 , surfactant 10 2:1 :1 surfactant 7, surfactant 10, surfactant 17 2:1 :1 surfactant 7, surfactant 13, surfactant 14, surfactant 17 1 : 1 : 1 : 1 surfactant 7, surfactant 8, surfactant 13, surfactant 15 1 : 1 : 1 : 1 surfactant 8 surfactant 8, surfactant 1 , surfactant 18 2:1 :1 surfactant 8, surfactant 17 1 :1 surfactant 8, surfactant 10, surfactant 11 , surfactant 12 1 : 1 : 1 : 1

[0053] GAR-P04610W010 Application (final). docx ABK

[0054] 13 surfactant 8, surfactant 11 , surfactant 14, surfactant 15 1 : 1 : 1 : 1 surfactant 8, surfactant 12, surfactant 14, surfactant 15 1 : 1 : 1 : 1 surfactant 8, surfactant 9, surfactant 10, surfactant 14 1 : 1 : 1 : 1 surfactant 9, surfactant 2, surfactant 4 2:1 :1

[0055] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer wherein formulation comprises the following diblock co-polymer surfactants and triblock co-polymer surfactants selected from surfactants molar ratio surfactant 1 surfactant 2, surfactant 4, surfactant 5, surfactant 12 1 :1 :1 :1 surfactant 2, surfactant 4, surfactant 8, surfactant 12 1 :1 :1 :1 surfactant 4, surfactant 5, surfactant 6 2:1 :1.

[0056] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the at least one amphiphilic polymer has a purity of at least 95% as determined by NMR spectroscopy.

[0057] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the at least one amphiphilic polymer has a purity of at least 98% as determined by NMR spectroscopy.

[0058] Definitions

[0059] Liposomes are composed of a lipid bilayer separating an aqueous internal compartment from the bulk aqueous phase. A liposome is a spherical vesicle having at least one lipid bilayer. The major types of liposomes are the multilamellar vesicles (MLV, with several lamellar phase lipid bilayers), the small unilamellar liposome vesicles (SUV, with one lipid bilayer), the large unilamellar vesicles (LUV), and the giant unilamellar vesicles (GUV).

[0060] Small unilamellar vesicles, large unilamellar vesicles and giant unilamellar vesicles are usually spherical with a diameter of typically 25 to 50 nm for small unilamellar vesicles, with a diameter of typically more than 50 to 1 ,000 nm for large unilamellar vesicles, and with a diameter of typically more than 1 to 1 ,000 pm for giant unilamellar vesicles. However, the chemical and mechanical instabilities of unsaturated fatty acids under high ionic strength conditions, especially multivalent cations, and their sensitivity to pH changes are considered to be the main challenges in utilizing these vesicles. In addition, inserting molecules into these vesicles represents a particular challenge given their impermeability and mechanical instability.

[0061] GAR-P04610W010 Application (final). docx The term “vesicle” refers to a non-natural or synthetic membranous and usually fluid- filled pouch resulting from the supramolecular assembly of lipids including, but not limited to, phospholipids. The interior contents of a phospholipid vesicle are separated from the exterior surroundings by at least one phospholipid bilayer. A phospholipid bilayer is a sheet of lipids two molecules thick, arranged so that the hydrophilic phosphate heads point “out” to the solution on either side of the bilayer and the hydrophobic tails point “in” to the core of the bilayer. This results in two “leaflets” which are each a single molecular layer of phospholipids. A “unilamellar vesicle” refers to a vesicle comprising only one phospholipid bilayer; “multilamellar vesicle” refers to a vesicle comprising more than one phospholipid bilayer. A “symmetric bilayer” is defined as a bilayer possessing two leaflets of the same composition, whereas an “asymmetric bilayer” is defined as a bilayer possessing two leaflets which differ in composition. The vesicle can have a unilamellar bilayer including a lipid and a component integrated by the bilayer. The integrated component can be a protein or a fragment thereof, a peptide, a polypeptide, an enzyme or a fragment thereof. Exemplary proteins are ATPase, E- cadherins, integrins, alpha-haemolisin. An exemplary polypeptide is the antibiotic gramicidin.

[0062] A "water-based droplet" is a droplet, which contains water or a dispersion of any substance in water. Also, a water-based droplet is a droplet, which consists of water or of a dispersion of any substance in water.

[0063] "Giant unilamellar vesicles" (“GUV”) are unilamellar vesicles, preferably spherical, and with a diameter comprised between 1 pm and 100 pm. The diameter of giant unilamellar vesicles is preferably measured by confocal microscopy. The term “giant” as used herein refers to unilamellar vesicles which form a lipid compartment on the mesoscale.

[0064] For GUV generation using w / o droplets, an aqueous solution containing SUVs is used to produce droplets in a fluorinated oil. To stabilize the water droplet against fusion, amphiphilic polymer surfactants are added to the oil phase, forming a surfactant layer on the water-oil interface. Due to charge-mediated interactions between the surfactants and the SUVs, a lipid bilayer self assembles and dsGUVs are formed. By destabilizing the droplets shell, the GUVs can be released. These droplets can either be produced by microfluidics, emulsification, or shaking. The formulations according to the present invention are particularly suitable for the droplet-based microfluidic method and are there particularly useful for droplet-based microfluidic applications.

[0065] GAR-P04610W010 Application (final). docx AOK

[0066] 15

[0067] The terms "cell delivery", "cell-targeting", "delivery", "cell type specific delivery" or "targeted delivery" are used interchangeably herein to refer to the delivery of particular agents or compounds or macromolecules by the giant unilamellar vesicles to regions, tissues or cells, intracellularly or extracellularly, in vivo or in vitro.

[0068] When the delivery occurs in the extracellular space of the cells, it means that the agent or compound or macromolecule is brought at a sufficient small distance to the target cell to allow interaction of the agent or compound or macromolecule with said cell, and thus the desired effect on said cell.

[0069] The agent or compound or macromolecule can be present on the surface of the giant unilamellar vesicles or in the lumen of giant unilamellar vesicles.

[0070] The terms “protein” and “peptide” are used interchangeably herein to refer to a polymer of amino acid residues. These terms apply to naturally occurring amino acid polymers and non-naturally occurring amino acid polymers, as well as to amino acid polymers in which one or more amino acid residues are an artificial chemical mimetic of a corresponding naturally occurring amino acid. As used herein, the terms encompass amino acid chains of any length, including full-length proteins, wherein the amino acid residues are linked by covalent peptide bonds, and fragments thereof.

[0071] The terms “nucleic acid”, “oligonucleotide,” and “polynucleotide” refer to deoxyribonucleic acids (DNA) or ribonucleic acids (RNA) and polymers thereof in either single- or double-stranded form. Unless specifically limited, the term encompasses nucleic acids containing known analogues of natural nucleotides that have similar binding properties as the reference nucleic acid and are metabolized in a manner similar to naturally occurring nucleotides.

[0072] The term “volatile” refers to organic solvents such as acetonitrile, nitromethane, piperidine, sulfolane, dimethylcarbonate, ethylencarbonate, dimethylsulfoxide, ether such as tetrahydrofurane (THF), diethylether, ketones such as acetone, butanone or pentanone, amides such as dimethylformamide (DMF) or dimethylacetamide, halogenated solvents such as chloroform, methylene chloride, tetrachloromethane, carboxylic acid ester such as acetic acid ethyl ester, alcohols such as methanol, ethanol, propanol, isopropanol, carboxylic acids such as formic acid, acetic acid, propionic acid, butyric acid hexyl ester, formamide, petrolether, ethers such as methyltert. -butylether (MTDC) or dioxane, hydrocarbons such as hexane, pentane or cyclohexane, aromatic solvents such as toluene, benzene, chlorobenzene, nitrobenzene, aniline, pyridine, anisol or xylene. Volatile organic solvents have preferably a boiling point below 215°C.

[0073] GAR-P04610W010 Application (final). docx “Aqueous mixture” or "aqueous phase" or "water phase" refers to a solution or suspension of lipids or other molecules substantially in water. The aqueous mixtures are immiscible with the oil-based mixtures of the present invention.

[0074] The term “water-immiscible fluorocarbon solvent” as used herein refers to non-polar fluidic chemical compounds with multiple carbon-fluorine bonds that are not miscible with water. Distinctive properties such as enhanced stability, low viscocity, low surface tension, low heats of vaporization, volatility, lipophobicity or hydrophobicity, arise due to the multiple C-F bonds. Said fluorocarbon solvents are preferably liquid at room temperature. The term “water-immiscible fluorocarbon solvents” include perfluorinated organofluorine compounds that lack a single C-H bond, as well as perfluorinated organofluorine compounds that substantially lack a C-H bond, as well as organofluorine compounds that partially comprise C-H bonds. Examplary compound classes of water- immiscible fluorocarbon solvents include, but are not restricted to, perfluorocarbons and hydrofluoroethers. Exemplary “water-immiscible fluorocarbon solvents”, synonymously used as “oils” herein, suitable for the preparation of certain lipid membranes and vesicles in the present invention include, but are not limited to, HFE- 7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1-ethoxy-1 ,1 ,2,2,3,3,4,4,4-nonafluorobutane and 1 -ethoxy-1 , 1 ,2, 3,3,3- hexafluoro-2-(trifluoromethyl)propane), HFE-7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3- methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 , 2, 3, 4, 4, 5, 5, 6,6,6- dodecafluoro-2-(trifluoromethyl)hexane), FC-40 (1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N,N- bis(nonafluorobutyl)butan-1 -amine), FC-43 (1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N, N- bis(1 ,1 ,2,2,3,3,4,4,4-nonafluorobutyl)butan-1-amine), FC-70 (1 ,1 , 2, 2, 3, 3, 4, 4, 5,5,5- undecafluoro-N,N-bis(undecafluoropentyl)pentan-1 -amine), FC-72

[0075] (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), or perfluorodecalin. One of skill in the art will appreciate that other solvents are useful as the oil mixture in the present invention.

[0076] The "polymer shell" of the giant unilamellar vesicles is made of an amphiphilic copolymer having a lipophilic end arranged at the outer side and a hydrophilic end arranged at the inner side of the polymer shell. This may be achieved by forming the polymer shell of the droplet from a diblock co-polymer or a triblock co-polymer.

[0077] The term “perfluorinated polymer” or “perfluorinated polymer block” refers to a polymeric organofluorine compound that lacks C-H bonds.

[0078] GAR-P04610W010 Application (final). docx The term “perfluorinated polyether”, “PFPE” or “perfluorinated polyther block” refers to a fluorpolymer consisting of a polymer chain in which monomers consisting of perfluoro-alkyl groups are joined by ether linkages, which has the structure: CF3-CF2-[O-C(CF3)(F)-CF2]n-O-C(CF3)(F)-COOH.

[0079] The general Formula of Krytox™ is: wherein n is an integer comprised between 10 and 60.

[0080] As used herein, the molecular weight values for the amphiphilic polymers, including the PEG (polyethylene glycol) and PFPE (perfluoropolyether) blocks, refer to the weight average molecular weight (Mw) unless otherwise specified. These molecular weights are determined by gel permeation chromatography (GPC) using appropriate calibration standards. For perfluorinated polyether blocks, GPC measurements are carried out using perfluoropolyether standards dissolved in a suitable fluorinated solvent (e.g., hexafluorobenzene or FC-75) and detected by refractive index. The PEG blocks are measured in analogous fashion using PEG standards.

[0081] For example, weight average molecular weight may be determined by GPC measurements using tetrahydrofuran (THF) as eluent and a refractive index detector. The system is calibrated with a series of polymer standards of narrow molecular weight distribution, covering a range from approximately 1 ,000 to 100,000 g / mol. A suitable column, such as a linear PLgel column (e.g., 5 pm, Mixed-C) can be used, with a flow rate of 1.0 mL / min at room temperature. Polymer samples are dissolved in THF and filtered prior to injection. An examplary sample volume of approximately 50 pL is injected for each measurement.

[0082] The indication of block size by nominal molecular weight (e.g., PFPE(7000), PEG(600)) follows standard nomenclature in the field of fluorinated surfactants and is supported by precise analytical methods as disclosed herein.

[0083] As used herein, molecular weight values given in parentheses, such as "PFPE(2000 g / mol)" or "PEG(600 g / mol)", refer to the weight average molecular weight (Mw) of the respective polymer blocks. The values are determined by standard analytical techniques (e.g. gel permeation chromatography (GPC)). The use of such nominal

[0084] GAR-P04610W010 Application (final). docx values is common in the field of block copolymer synthesis and allows precise formulation control based on well-defined feedstock materials.

[0085] The molecular weights (Mw) derived from the GPC traces are then calculated relative to the polystyrene calibration curve. Optionally, universal calibration correction (e.g., using Mark-Houwink parameters) can be applied. Due to the structural differences between perfluoropolyether and polystyrene, the values obtained are relative molecular weights, but represent a consistent and widely accepted means of characterizing polymer block sizes in the field of fluorinated surfactants.

[0086] The term "monodisperse" or "monodispersed" refers to vesicles or droplets of uniform size in a dispersed phase, where the vesicles or droplets are present in an unaggregate and discrete state. The dispersed phase according to the invention is preferably a water phase for the polymer shell stabilized giant unilamellar vesicles. “Dispersed phase” is the droplet phase whereas continuous phase would be the surrounding oil, volatile solvent, or even air before droplets are collected into solution.

[0087] Vesicles of "uniform size" means that the polymer shell stabilized giant unilamellar vesicles obtained after splitting, as well as the giant unilamellar vesicles released from the polymer shell, show a coefficient of variation in size lower than 16%, lower than 15%, lower than 14%, lower than 13%, lower than 12%, lower than 11%, lower than 10%.

[0088] The (average) molecular weight of the polymers and polymer blocks described herein, is determined with gel permeation chromatography using a polystyrene standard.

[0089] Description of the invention

[0090] The present invention is directed to a formulation comprising a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant of formula A-B and a triblock co-polymer surfactant of formula B’-A’-B’, wherein A and A’ independently of each other represent a water-soluble polymer block having a range in molecular weight from 300 g / mol to 2000 g / mol, and B and B’ represent independently of each other a perfluorinated polyether block having a range in molecular weight from 1500 g / mol to 7500 g / mol, wherein the weight average molecular weights of A, A’, B, and B’ are determined by gel permeation chromatography, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9 have the following formula:

[0091] GAR-P04610W010 Application (final). docx ABK surfactant 1 : PFPE(2000 g / mol-L1-PEG(350 g / mol) surfactant 2: PFPE(4000 g / mol)-L1-PEG(350 g / mol) surfactant 3: PFPE(7000 g / mol)-L1-PEG(350 g / mol) surfactant 4: PFPE(2000 g / mol)-L1-PEG(550 g / mol) surfactant 5: PFPE(4000 g / mol)-L1-PEG(550 g / mol) surfactant 6: PFPE(7000 g / mol)-L1-PEG(550 g / mol) surfactant 7: PFPE(2000 g / mol)-L1-PEG(750 g / mol) surfactant 8: PFPE(4000 g / mol)-L1-PEG(750 g / mol) surfactant 9: PFPE(7000 g / mol)-L1-PEG(750 g / mol) wherein L1represents a triazole ring having the structure: and wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein surfactant 10 to surfactant 18 have the following formula: surfactant 10: PFPE(2000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(2000 g / mol) surfactant 11 : PFPE(4000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(4000 g / mol) surfactant 12: PFPE(7000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(7000 g / mol) surfactant 13: PFPE(2000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(2000 g / mol) surfactant 14: PFPE(4000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(4000 g / mol) surfactant 15: PFPE(7000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(7000 g / mol) surfactant 16: PFPE(2000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(2000 g / mol) surfactant 17: PFPE(4000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(4000 g / mol) surfactant 18: PFPE(7000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(7000 g / mol) wherein L2and L3represent a triazole ring having the structures: Surfactant 1 to surfactant 9 have the following formula, wherein “L1” represents a triazole ring having the structure:

[0092] GAR-P04610W010 Application (final). docx ABK

[0093] 20

[0094] Surfactant 1 PFPE(2000 g / mol-L1-PEG(350 g / mol)

[0095] Surfactant 2 PFPE(4000 g / mol)-L1-PEG(350 g / mol)

[0096] Surfactant 3 PFPE(7000 g / mol)-L1-PEG(350 g / mol)

[0097] Surfactant 4 PFPE(2000 g / mol)-L1-PEG(550 g / mol)

[0098] Surfactant 5 PFPE(4000 g / mol)-L1-PEG(550 g / mol)

[0099] Surfactant 6 PFPE(7000 g / mol)-L1-PEG(550 g / mol),

[0100] Surfactant 7 PFPE(2000 g / mol)-L1-PEG(750 g / mol),

[0101] Surfactant 8 PFPE(4000 g / mol)-L1-PEG(750 g / mol),

[0102] Surfactant 9 PFPE(7000 g / mol)-L1-PEG(750 g / mol).

[0103] Surfactant 10 to surfactant 18 have the following formula, wherein “L2” and “L3” represent a triazole ring having the structures:

[0104] Surfactant 10 PFPE(2000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(2000 g / mol),

[0105] Surfactant 11 PFPE(4000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(4000 g / mol),

[0106] Surfactant 12 PFPE(7000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(7000 g / mol),

[0107] Surfactant 13 PFPE(2000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(2000 g / mol),

[0108] Surfactant 14 PFPE(4000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(4000 g / mol),

[0109] Surfactant 15 PFPE(7000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(7000 g / mol),

[0110] Surfactant 16 PFPE(2000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(2000 g / mol),

[0111] Surfactant 17 PFPE(4000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(4000 g / mol),

[0112] Surfactant 18 PFPE(7000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(7000 g / mol).

[0113] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and one amphiphilic polymer selected from surfactant 1 to surfactant 18.

[0114] Preferably, the concentration of the amphiphilic polymer is in the range between 1 mM and 20 mM, preferably between 4 mM and 6 mM and most preferably 5 mM, when the amphiphilic polymer is selected from surfactant 1 to 9. Preferably, the concentration of the amphiphilic polymer is in the range between 0.5 mM and 10 mM, preferably

[0115] GAR-P04610W010 Application (final). docx between 1.5 mM and 3.5 mM and most preferably 2.5 mM, when the amphiphilic polymer is selected from surfactant 10 to 18.

[0116] Preferably, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2, 2, 3, 3, 4, 4- nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2- (trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 -ethoxy-1 , 1 ,2, 2, 3, 3, 4,4,4- nonafluorobutane and 1 -ethoxy-1 , 1 ,2, 3, 3, 3-hexafluoro-2-(trifluoromethyl)propane), HFE- 7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3-methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), FC-40

[0117] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), FC-43

[0118] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(1 ,1 ,2,2,3, 3,4,4,4-nonafluorobutyl)butan-1 -amine), FC-70 (1 ,1 ,2,2,3,3,4,4,5,5,5-undecafluoro-N,N-bis(undecafluoropentyl)pentan-1-amine), FC-72 (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned.

[0119] More preferably, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3, 3,4,4-nonafluoro-4-methoxy- butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7500 (3- ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), and FC-40 (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine).

[0120] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and one amphiphilic polymer selected from surfactant 1 to surfactant 18, wherein the formulation is devoid of free perfluoropolyether carboxylic acid.

[0121] In another preferred embodiment, the inventive formulation comprises a water- immiscible fluorocarbon solvent, one amphiphilic polymer selected from surfactant 1 to surfactant 18, and 0.1 mM to 15 mM of a perfluoropolyether carboxylic acid. Preferably, the perfluoropolyether carboxylic acid is a polymer of hexafluoropropylene oxide.

[0122] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and two amphiphilic polymers independently selected from surfactant 1 to surfactant 18.

[0123] Preferably, the molar ratio of the two amphiphilic polymers is 3: 1 , 1 :1 or 1 :3. Preferably, the total concentration of the two amphiphilic polymers is in a range between 0.5 mM x a + 0.5 mM and 10 mM x a + 10 mM, wherein a is the molar fraction of the diblock copolymer surfactant in the two amphiphilic polymers.

[0124] GAR-P04610W010 Application (final). docx More preferably, the total concentration of the two amphiphilic polymers is 2.5 mM x a + 2.5 mM, wherein a is the molar fraction of the diblock co-polymer surfactant in the two amphiphilic polymers.

[0125] Preferably, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2, 2, 3, 3, 4, 4- nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2- (trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 -ethoxy-1 , 1 ,2, 2, 3, 3, 4,4,4- nonafluorobutane and 1 -ethoxy-1 , 1 ,2, 3, 3, 3-hexafluoro-2-(trifluoromethyl)propane), HFE- 7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3-methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), FC-40

[0126] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), FC-43

[0127] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(1 ,1 ,2,2,3, 3,4,4,4-nonafluorobutyl)butan-1 -amine), FC-70 (1 ,1 ,2,2,3,3,4,4,5,5,5-undecafluoro-N,N-bis(undecafluoropentyl)pentan-1-amine), FC-72 (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned.

[0128] More preferably, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3, 3,4,4-nonafluoro-4-methoxy- butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7500 (3- ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), and FC-40 (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine).

[0129] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and two amphiphilic polymers independently selected from surfactant 1 to surfactant 18, wherein the formulation is devoid of free perfluoropolyether carboxylic acid.

[0130] In another preferred embodiment, the inventive formulation comprises a water- immiscible fluorocarbon solvent, two amphiphilic polymers independently from surfactant 1 to surfactant 18, and 0.1 mM to 15 mM of a perfluoropolyether carboxylic acid. Preferably, the perfluoropolyether carboxylic acid is a polymer of hexafluoropropylene oxide.

[0131] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and three amphiphilic polymers independently selected from surfactant 1 to surfactant 18.

[0132] Preferably, the molar ratio of the three amphiphilic polymers is 2:1 :1 , 1 :2:1 , 1 :1 :1 or 1 :1 :2. Preferably, the total concentration of the three amphiphilic polymers is in a range

[0133] GAR-P04610W010 Application (final). docx between 0.5 mM x a + 0.5 mM and 10 mM x a + 10 mM, wherein a is the molar fraction of the diblock co-polymer surfactant in the three amphiphilic polymers.

[0134] More preferably, the total concentration of the two amphiphilic polymers is 2.5 mM x a + 2.5 mM, wherein a is the molar fraction of the diblock co-polymer surfactant in the three amphiphilic polymers.

[0135] Preferably, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2, 2, 3, 3, 4, 4- nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2- (trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 -ethoxy-1 , 1 ,2, 2, 3, 3, 4,4,4- nonafluorobutane and 1 -ethoxy-1 , 1 ,2, 3, 3, 3-hexafluoro-2-(trifluoromethyl)propane), HFE- 7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3-methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), FC-40

[0136] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), FC-43

[0137] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(1 ,1 ,2,2,3, 3,4,4,4-nonafluorobutyl)butan-1 -amine), FC-70 (1 ,1 ,2,2,3,3,4,4,5,5,5-undecafluoro-N,N-bis(undecafluoropentyl)pentan-1-amine), FC-72 (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned.

[0138] More preferably, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3, 3,4,4-nonafluoro-4-methoxy- butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7500 (3- ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), and FC-40 (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine).

[0139] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and three amphiphilic polymers independently selected from surfactant 1 to surfactant 18, wherein the formulation is devoid of free perfluoropolyether carboxylic acid.

[0140] In another preferred embodiment, the inventive formulation comprises a water- immiscible fluorocarbon solvent, three amphiphilic polymers independently from surfactant 1 to surfactant 18, and 0.1 mM to 15 mM of a perfluoropolyether carboxylic acid. Preferably, the perfluoropolyether carboxylic acid is a polymer of hexafluoropropylene oxide.

[0141] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and four amphiphilic polymers independently selected from surfactant 1 to surfactant 18.

[0142] GAR-P04610W010 Application (final). docx Preferably, the molar ratio of the four amphiphilic polymers is 1 :1 :1 :1 . Preferably, the total concentration of the four amphiphilic polymers is in a range between 0.5 mM x a + 0.5 mM and 10 mM x a + 10 mM, wherein a is the molar fraction of the diblock copolymer surfactant in the four amphiphilic polymers.

[0143] More preferably, the total concentration of the two amphiphilic polymers is 2.5 mM x a + 2.5 mM, wherein a is the molar fraction of the diblock co-polymer surfactant in the four amphiphilic polymers.

[0144] Preferably, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 , 2, 2, 3, 3,4,4- nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2- (trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 -ethoxy-1 , 1 ,2, 2, 3, 3, 4,4,4- nonafluorobutane and 1 -ethoxy-1 , 1 ,2, 3, 3, 3-hexafluoro-2-(trifluoromethyl)propane), HFE- 7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3-methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), FC-40

[0145] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), FC-43

[0146] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(1 ,1 ,2,2,3, 3,4,4,4-nonafluorobutyl)butan-1 -amine), FC-70 (1 ,1 ,2,2,3,3,4,4,5,5,5-undecafluoro-N,N-bis(undecafluoropentyl)pentan-1-amine), FC-72 (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned.

[0147] More preferably, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3, 3,4,4-nonafluoro-4-methoxy- butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7500 (3- ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), and FC-40 (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine).

[0148] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and four amphiphilic polymers independently selected from surfactant 1 to surfactant 18, wherein the formulation is devoid of free perfluoropolyether carboxylic acid.

[0149] In another preferred embodiment, the inventive formulation comprises a water- immiscible fluorocarbon solvent, four amphiphilic polymers independently from surfactant 1 to surfactant 18, and 0.1 mM to 15 mM of a perfluoropolyether carboxylic acid. Preferably, the perfluoropolyether carboxylic acid is a polymer of hexafluoropropylene oxide.

[0150] GAR-P04610W010 Application (final). docx Preferably, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2, 2, 3, 3, 4, 4- nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2- (trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 -ethoxy-1 , 1 ,2, 2, 3, 3, 4,4,4- nonafluorobutane and 1 -ethoxy-1 , 1 ,2, 3, 3, 3-hexafluoro-2-(trifluoromethyl)propane), HFE- 7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3-methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), FC-40

[0151] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), FC-43

[0152] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(1 ,1 ,2,2,3, 3,4,4,4-nonafluorobutyl)butan-1 -amine), FC-70 (1 ,1 ,2,2,3,3,4,4,5,5,5-undecafluoro-N,N-bis(undecafluoropentyl)pentan-1-amine), FC-72 (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned.

[0153] More preferably, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 , 1 ,1 , 2,3,3- hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 - ethoxy-1 , 1 ,2,2,3,3,4,4,4-nonafluorobutane and 1 -ethoxy-1 , 1 , 2,3,3, 3-hexafluoro-2- (trifluoromethyl)propane), HFE-7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3-methoxy-4-

[0154] (trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2- (trifluoromethyl)hexane), FC-40 (1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N,N- bis(nonafluorobutyl)butan-1 -amine), perfluorodecalin, or mixtures of the aforementioned.

[0155] More preferably, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3, 3,4,4-nonafluoro-4-methoxy- butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7500 (3- ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), and FC-40 (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine).

[0156] Preferably, the inventive formulation is devoid of free perfluoropolyether carboxylic acid.

[0157] Preferably, the inventive formulation further comprises 0.1 mM to 15 mM of a perfluoropolyether carboxylic acid. Preferably, said perfluoropolyether carboxylic acid is a polymer of hexafluoropropylene oxide.

[0158] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one

[0159] GAR-P04610W010 Application (final). docx amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock copolymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the total concentration of the at least one amphiphilic polymer is in the range between 0.5 mM x a + 0.5 mM and 10 mM x a + 10 mM, more preferably between 2.5 mM x a + 1.5 mM and 2.5 mM x a + 3.5 mM, most preferably 2.5 mM x a + 2.5 mM.

[0160] The coefficient a as used herein is the molar fraction of the diblock co-polymer surfactant(s) in the total amphiphilic polymers and ranges between 0 and 1 .

[0161] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock copolymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, and wherein the water-immiscible fluorocarbon solvent is selected from the group comprising or consisting of HFE-7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 , 2, 2, 3, 3,4,4- nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2- (trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 -ethoxy-1 , 1 ,2, 2, 3, 3, 4,4,4- nonafluorobutane and 1 -ethoxy-1 , 1 ,2, 3, 3, 3-hexafluoro-2-(trifluoromethyl)propane), HFE- 7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3-methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), FC-40

[0162] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), FC-43

[0163] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(1 ,1 ,2,2,3, 3,4,4,4-nonafluorobutyl)butan-1 -amine), FC-70 (1 ,1 ,2,2,3,3,4,4,5,5,5-undecafluoro-N,N-bis(undecafluoropentyl)pentan-1-amine), FC-72 (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned.

[0164] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-

[0165] GAR-P04610W010 Application (final). docx polymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, and wherein the water-immiscible fluorocarbon solvent is selected from the group comprising or consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 , 1 ,1 , 2,3,3- hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7500 (3-ethoxy- 1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), and FC-40 (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine).

[0166] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock copolymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, and wherein the water-immiscible fluorocarbon solvent is selected from the group comprising or consisting of HFE-7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 , 2, 2, 3, 3,4,4- nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2- (trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 -ethoxy-1 , 1 ,2, 2, 3, 3, 4,4,4- nonafluorobutane and 1 -ethoxy-1 , 1 ,2, 3, 3, 3-hexafluoro-2-(trifluoromethyl)propane), HFE- 7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3-methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), FC-40

[0167] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), FC-43

[0168] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(1 ,1 ,2,2,3, 3,4,4,4-nonafluorobutyl)butan-1 -amine), FC-70 (1 ,1 ,2,2,3,3,4,4,5,5,5-undecafluoro-N,N-bis(undecafluoropentyl)pentan-1-amine), FC-72 (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned.

[0169] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock copolymer surfactant, wherein the diblock co-polymer surfactant is selected from the

[0170] GAR-P04610W010 Application (final). docx group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, and wherein the water-immiscible fluorocarbon solvent is selected from the group comprising or consisting of HFE-7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 , 2, 2, 3, 3,4,4- nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2- (trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 -ethoxy-1 , 1 ,2, 2, 3, 3, 4,4,4- nonafluorobutane and 1 -ethoxy-1 , 1 ,2, 3, 3, 3-hexafluoro-2-(trifluoromethyl)propane), HFE- 7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3-methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), FC-40

[0171] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), FC-43

[0172] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(1 ,1 ,2,2,3, 3,4,4,4-nonafluorobutyl)butan-1 -amine), FC-70 (1 ,1 ,2,2,3,3,4,4,5,5,5-undecafluoro-N,N-bis(undecafluoropentyl)pentan-1-amine), FC-72 (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned.

[0173] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent the following diblock co-polymer surfactants and triblock co-polymer surfactants selected from, surfactant 1 surfactant 1 , surfactant 10, surfactant 11 surfactant 1 , surfactant 10, surfactant 17 surfactant 1 , surfactant 11 , surfactant 13, surfactant 15 surfactant 1 , surfactant 13, surfactant 15, surfactant 16 surfactant 1 , surfactant 2, surfactant 4, surfactant 15 surfactant 1 , surfactant 3, surfactant 7, surfactant 11 surfactant 1 , surfactant 4, surfactant 11 , surfactant 13 surfactant 1 , surfactant 4, surfactant 11 , surfactant 16 surfactant 1 , surfactant 4, surfactant 18 surfactant 1 , surfactant 6, surfactant 11 , surfactant 12 surfactant 1 , surfactant 7, surfactant 11 , surfactant 12 surfactant 1 , surfactant 7, surfactant 13, surfactant 14 surfactant 1 , surfactant 8, surfactant 15, surfactant 18 surfactant 1 , surfactant 9, surfactant 13, surfactant 16 surfactant 10 surfactant 10, surfactant 13, surfactant 14, surfactant 15 surfactant 10, surfactant 14, surfactant 17, surfactant 18 surfactant 11 surfactant 11 , surfactant 6, surfactant 12 surfactant 11 , surfactant 9, surfactant 16 surfactant 12 surfactant 12, surfactant 13, surfactant 17

[0174] GAR-P04610W010 Application (final). docx ABK

[0175] 29 surfactant 13 surfactant 13, surfactant 14, surfactant 18 surfactant 13, surfactant 15 surfactant 13, surfactant 7, surfactant 9 surfactant 14 surfactant 14, surfactant 10, surfactant 17 surfactant 14, surfactant 3, surfactant 15 surfactant 15 surfactant 15, surfactant 12, surfactant 14 surfactant 15, surfactant 14, surfactant 18 surfactant 16 surfactant 16, surfactant 14 surfactant 16, surfactant 8, surfactant 18 surfactant 17 surfactant 17, surfactant 5, surfactant 13 surfactant 17, surfactant 8, surfactant 18 surfactant 17, surfactant 8, surfactant 9 surfactant 18 surfactant 18, surfactant 1 , surfactant 13 surfactant 18, surfactant 10, surfactant 13 surfactant 18, surfactant 14, surfactant 16 surfactant 18, surfactant 5, surfactant 11 surfactant 18, surfactant 5, surfactant 12 surfactant 18, surfactant 9, surfactant 13 surfactant 2, surfactant 11 , surfactant 18 surfactant 2, surfactant 3, surfactant 4, surfactant 10 surfactant 2, surfactant 3, surfactant 4, surfactant 12 surfactant 2, surfactant 3, surfactant 8 surfactant 2, surfactant 4, surfactant 5, surfactant 12 surfactant 2, surfactant 4, surfactant 8, surfactant 12 surfactant 2, surfactant 5, surfactant 18 surfactant 2, surfactant 6, surfactant 15, surfactant 16 surfactant 3, surfactant 1 , surfactant 15 surfactant 3, surfactant 10, surfactant 11 surfactant 3, surfactant 10, surfactant 12, surfactant 16 surfactant 3, surfactant 15, surfactant 16 surfactant 3, surfactant 2, surfactant 7 surfactant 3, surfactant 4, surfactant 10, surfactant 13 surfactant 3, surfactant 4, surfactant 16 surfactant 3, surfactant 4, surfactant 5, surfactant 16 surfactant 3, surfactant 4, surfactant 8 surfactant 3, surfactant 7, surfactant 15 surfactant 3, surfactant 8, surfactant 14, surfactant 16 surfactant 4, surfactant 5, surfactant 6 surfactant 5, surfactant 12, surfactant 16 surfactant 5, surfactant 13, surfactant 17 surfactant 5, surfactant 14 surfactant 5, surfactant 6, surfactant 11 , surfactant 15 surfactant 5, surfactant 7, surfactant 12, surfactant 14 surfactant 6 surfactant 6, surfactant 13, surfactant 17 surfactant 6, surfactant 15

[0176] GAR-P04610W010 Application (final). docx AOK

[0177] 30 surfactant 7 surfactant 7, surfactant 1 surfactant 7, surfactant 1 , surfactant 10 surfactant 7, surfactant 10, surfactant 17 surfactant 7, surfactant 11 , surfactant 12, surfactant 16 surfactant 7, surfactant 12, surfactant 13, surfactant 17 surfactant 7, surfactant 13, surfactant 14, surfactant 17 surfactant 7, surfactant 16 surfactant 7, surfactant 8, surfactant 13, surfactant 15 surfactant 8 surfactant 8, surfactant 1 , surfactant 18 surfactant 8, surfactant 10, surfactant 11 , surfactant 12 surfactant 8, surfactant 11 , surfactant 14, surfactant 15 surfactant 8, surfactant 12, surfactant 14, surfactant 15 surfactant 8, surfactant 17 surfactant 8, surfactant 3, surfactant 15 surfactant 8, surfactant 9, surfactant 10, surfactant 14 surfactant 9 surfactant 9, surfactant 11 , surfactant 12, surfactant 16 surfactant 9, surfactant 17 surfactant 9, surfactant 18 surfactant 9, surfactant 2, surfactant 4. wherein the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7500 (3-ethoxy- 1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), and FC-40

[0178] (1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1-amine).

[0179] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent the following diblock co-polymer surfactants and triblock co-polymer surfactants selected from surfactants molar ratio surfactant 1 surfactant 1 , surfactant 10, surfactant 11 2:1 :1 surfactant 1 , surfactant 10, surfactant 17 2:1 :1 surfactant 1 , surfactant 2, surfactant 4, surfactant 15 1 : 1 : 1 : 1 surfactant 1 , surfactant 3, surfactant 7, surfactant 11 1 : 1 : 1 : 1 surfactant 1 , surfactant 4, surfactant 11 , surfactant 13 1 : 1 : 1 : 1 surfactant 1 , surfactant 4, surfactant 11 , surfactant 16 1 : 1 : 1 : 1 surfactant 1 , surfactant 4, surfactant 18 2:1 :1 surfactant 1 , surfactant 6, surfactant 11 , surfactant 12 1 : 1 : 1 : 1 surfactant 1 , surfactant 7, surfactant 11 , surfactant 12 1 : 1 : 1 : 1 surfactant 1 , surfactant 7, surfactant 13, surfactant 14 1 : 1 : 1 : 1 surfactant 1 , surfactant 8, surfactant 15, surfactant 18 1 : 1 : 1 : 1 surfactant 1 , surfactant 9, surfactant 13, surfactant 16 1 : 1 : 1 : 1 surfactant 11 , surfactant 6, surfactant 12 2:1 :1 surfactant 13

[0180] GAR-P04610W010 Application (final). docx ABK

[0181] 31 surfactant 13, surfactant 7, surfactant 9 2:1 :1 surfactant 14, surfactant 3, surfactant 15 2:1 :1 surfactant 17, surfactant 5, surfactant 13 2:1 :1 surfactant 17, surfactant 8, surfactant 9 2:1 :1 surfactant 18, surfactant 5, surfactant 11 2:1 :1 surfactant 18, surfactant 5, surfactant 12 2:1 :1 surfactant 2, surfactant 11 , surfactant 18 2:1 :1 surfactant 2, surfactant 3, surfactant 4, surfactant 10 1 : 1 : 1 : 1 surfactant 2, surfactant 3, surfactant 4, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 3, surfactant 8 2:1 :1 surfactant 2, surfactant 4, surfactant 5, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 4, surfactant 8, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 5, surfactant 18 2:1 :1 surfactant 2, surfactant 6, surfactant 15, surfactant 16 1 : 1 : 1 : 1 surfactant 3, surfactant 1 , surfactant 15 2:1 :1 surfactant 3, surfactant 4, surfactant 10, surfactant 13 1 : 1 : 1 : 1 surfactant 3, surfactant 4, surfactant 5, surfactant 16 1 : 1 : 1 : 1 surfactant 3, surfactant 4, surfactant 8 2:1 :1 surfactant 3, surfactant 7, surfactant 15 2:1 :1 surfactant 3, surfactant 8, surfactant 14, surfactant 16 1 : 1 : 1 : 1 surfactant 4, surfactant 5, surfactant 6 2:1 :1 surfactant 5, surfactant 12, surfactant 16 2:1 :1 surfactant 5, surfactant 13, surfactant 17 2:1 :1 surfactant 5, surfactant 14 1 :1 surfactant 5, surfactant 6, surfactant 11 , surfactant 15 1 : 1 : 1 : 1 surfactant 5, surfactant 7, surfactant 12, surfactant 14 1 : 1 : 1 : 1 surfactant 6 surfactant 6, surfactant 13, surfactant 17 2:1 :1 surfactant 6, surfactant 15 1 :1 surfactant 7 surfactant 7, surfactant 1 3:1 surfactant 7, surfactant 1 , surfactant 10 2:1 :1 surfactant 7, surfactant 10, surfactant 17 2:1 :1 surfactant 7, surfactant 13, surfactant 14, surfactant 17 1 : 1 : 1 : 1 surfactant 7, surfactant 8, surfactant 13, surfactant 15 1 : 1 : 1 : 1 surfactant 8 surfactant 8, surfactant 1 , surfactant 18 2:1 :1 surfactant 8, surfactant 17 1 :1 surfactant 8, surfactant 10, surfactant 11 , surfactant 12 1 : 1 : 1 : 1 surfactant 8, surfactant 11 , surfactant 14, surfactant 15 1 : 1 : 1 : 1 surfactant 8, surfactant 12, surfactant 14, surfactant 15 1 : 1 : 1 : 1 surfactant 8, surfactant 9, surfactant 10, surfactant 14 1 : 1 : 1 : 1 surfactant 9, surfactant 2, surfactant 4 2:1 :1 wherein the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7500 (3-ethoxy-

[0182] GAR-P04610W010 Application (final). docx 1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), and FC-40 (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine).

[0183] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent the following diblock co-polymer surfactants and triblock co-polymer surfactants selected from, surfactants molar ratio surfactant 1 surfactant 2, surfactant 4, surfactant 5, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 4, surfactant 8, surfactant 12 1 : 1 : 1 : 1 surfactant 4, surfactant 5, surfactant 6 2:1 :1. wherein the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7500 (3-ethoxy- 1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), and FC-40 (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine)

[0184] Likewise, the term “wherein the water-soluble polymer blocks and the perfluorinated polymer blocks are connected to each other by a triazole ring”, i.e. L1, L2or L3is understood that in the formulae A-B and B’-A’-B’, each represents a triazole ring. Thus, the the term “wherein the water-soluble polymer blocks and the perfluorinated polymer blocks are connected to each other by a triazole ring” does not mean that all water-soluble polymer blocks and all perfluorinated polymer blocks are connected to the same triazole ring.

[0185] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock copolymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the formulation is devoid of free perfluoropolyether carboxylic acid, and wherein the perfluoropolyether carboxylic acid is a polymer of hexafluoropropylene oxide.

[0186] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one

[0187] GAR-P04610W010 Application (final). docx amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock copolymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the water-immiscible fluorocarbon solvent is selected from the group comprising or consisting of HFE-7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE- 7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1-ethoxy-1 ,1 ,2,2,3,3,4,4,4-nonafluorobutane and 1 -ethoxy-1 , 1 ,2, 3,3,3- hexafluoro-2-(trifluoromethyl)propane), HFE-7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3- methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 , 2, 3, 4, 4, 5, 5, 6,6,6- dodecafluoro-2-(trifluoromethyl)hexane), FC-40 (1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N,N- bis(nonafluorobutyl)butan-1 -amine), FC-43 (1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N, N- bis(1 ,1 ,2,2,3,3,4,4,4-nonafluorobutyl)butan-1-amine), FC-70 (1 ,1 , 2, 2, 3, 3, 4, 4, 5,5,5- undecafluoro-N,N-bis(undecafluoropentyl)pentan-1 -amine), FC-72

[0188] (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned, wherein the formulation is devoid of free perfluoropolyether carboxylic acid, and wherein the perfluoropolyether carboxylic acid is a polymer of hexafluoropropylene oxide.

[0189] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock copolymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the formulation further comprises 0.1 mM to 15 mM of a perfluoropolyether carboxylic acid, and wherein the perfluoropolyether carboxylic acid is a polymer of hexafluoropropylene oxide.

[0190] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock copolymer surfactant, wherein the diblock co-polymer surfactant is selected from the

[0191] GAR-P04610W010 Application (final). docx group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9; wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the water-immiscible fluorocarbon solvent is selected from the group comprising or consisting of HFE-7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 , 2, 2, 3, 3,4,4- nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2- (trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 -ethoxy-1 , 1 ,2, 2, 3, 3, 4,4,4- nonafluorobutane and 1 -ethoxy-1 , 1 ,2, 3, 3, 3-hexafluoro-2-(trifluoromethyl)propane), HFE- 7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3-methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), FC-40

[0192] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), FC-43

[0193] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(1 ,1 ,2,2,3, 3,4,4,4-nonafluorobutyl)butan-1 -amine), FC-70 (1 ,1 ,2,2,3,3,4,4,5,5,5-undecafluoro-N,N-bis(undecafluoropentyl)pentan-1-amine), FC-72 (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned, wherein the formulation further comprises 0.1 mM to 15 mM of a perfluoropolyether carboxylic acid, and wherein the perfluoropolyether carboxylic acid is a polymer of hexafluoropropylene oxide.

[0194] The development of a fluorinated block copolymer surfactant, primarily utilized as diblock and triblock surfactants comprising polyethylene glycol (PEG) and perfluoropolyether (PFPE) has already been described in the prior art. The synthesis of these surfactants necessitates the combination of PEG and PFPE, with three distinct synthetic methodologies being highlighted: esterification, amidation, and copper(l)- catalyzed azide-alkyne cycloaddition (CuAAC,). Krytox, a PFPE variant with a terminal carboxylic acid functional group, serves as the foundational material for the PFPE segment.

[0195] The initial approach for merging PEG with Krytox™ involves esterification, employing n- butyllithium (n-BuLi) as a catalyst. This process begins with the deprotonation of PEG by n-BuLi under an inert atmosphere, resulting in lithiated PEG. Upon introducing Krytox to this environment, the lithiated oxygen atom of PEG attacks the carboxyl carbon of Krytox™, leading to the elimination of a hydroxide ion — a reaction facilitated by the basic conditions. This sequence culminates in the formation of the esterified triblock surfactant.

[0196] An alternative synthesis route involves the amidation of diamine PEG and Krytox™. This process is initiated by converting the carboxylic acid group of Krytox into an acid chloride using oxalyl chloride, again under inert conditions. The enhanced reactivity of

[0197] GAR-P04610W010 Application (final). docx the acid chloride enables its interaction with the amine groups of diamine PEG, resulting in the formation of the amidated triblock surfactant.

[0198] Both aforementioned synthetic strategies rely on the generation of a reactive, unstable intermediate that reacts with the counterpart polymer. However, the high reactivity and sluggish reaction kinetics, attributed to the substantial molecular weights involved, may lead to side reactions that reduce synthesis efficiency. To mitigate such issues, employing stable reactants is advisable. A prime example of this approach is the CuAAC click reaction, which involves the combination of diazide PEG and propargyl PFPE in the presence of copper(l) ions. These ions facilitate the 1 ,3-dipolar cycloaddition, allowing for the straightforward separation of the resultant clicked surfactant upon reaction completion.

[0199] Krytox™, when utilized as a precursor, may persist as a contaminant within the end product. Owing to its carboxylic acid moiety, it induces a reduction in the pH value and fosters the formation of a charged interface, both of which adversely affect the encapsulated material. Schrdter (in “Development of next-generation fluorinated block copolymers and bottom-up assembly of biomimetic cell interaction modules”, 2022, PhD thesis) demonstrated that the CuAAC reaction exhibits a lower residual Krytox™ content relative to the alternative pathways investigated.

[0200] In contemporary implementations of droplet-based microfluidics, a singular surfactant, characterized by the molecular weights of the employed polymers, is utilized within experimental frameworks. The molecular weight and purity of the aforementioned surfactant are contingent upon the nature of the starting material, the distributor, and the synthetic route employed.

[0201] In addition to the choice of synthetic pathway, the purity of the resulting amphiphilic polymer surfactant plays a decisive role in the performance of the formulation. Higher surfactant purity, for example at least 95 mol % as determined by1H NMR spectroscopy or13F NMR spectroscopy, reduces the presence of unreacted starting materials, side products, and low-molecular-weight fractions. Such impurities may otherwise destabilize droplet interfaces, interfere with encapsulated reagents, or introduce undesired chemical functionalities. Surfactants of higher purity exhibit improved batch-to-batch reproducibility, enhanced biocompatibility, and greater stability under microfluidic operating conditions. The removal of residual Krytox™ or other PFPE-based precursors in particular has been found to improve pH stability and minimize non-specific interactions at the droplet interface, thereby enhancing the reliability of droplet-based assays and encapsulation processes.

[0202] GAR-P04610W010 Application (final). docx For the purposes of the present application, the term “purity” refers to the molar percentage of the target amphiphilic polymer surfactant relative to the total molar amount of surfactant-related species present in the formulation, as determined by quantitative nuclear magnetic resonance (1H NMR and / or19F NMR) spectroscopy. Surfactant-related species include the desired amphiphilic polymer surfactant and any residual starting materials, unreacted precursors (e.g., PEG or PFPE derivatives), side products, degradation products, or low-molecular-weight homologues arising from the synthesis. The molar percentage is calculated by integrating characteristic NMR signals corresponding to the target amphiphilic polymer and comparing these to the combined integral of all surfactant-related species present.

[0203] Thus, the present invention is also directed to a formulation comprising a water- immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9 have the following formula: surfactant 1 : PFPE(2000 g / mol-L1-PEG(350 g / mol) surfactant 2: PFPE(4000 g / mol)-L1-PEG(350 g / mol) surfactant 3: PFPE(7000 g / mol)-L1-PEG(350 g / mol) surfactant 4: PFPE(2000 g / mol)-L1-PEG(550 g / mol) surfactant 5: PFPE(4000 g / mol)-L1-PEG(550 g / mol) surfactant 6: PFPE(7000 g / mol)-L1-PEG(550 g / mol) surfactant 7: PFPE(2000 g / mol)-L1-PEG(750 g / mol) surfactant 8: PFPE(4000 g / mol)-L1-PEG(750 g / mol) surfactant 9: PFPE(7000 g / mol)-L1-PEG(750 g / mol) wherein L1represents a triazole ring having the structure:

[0204] O NxN

[0205] .

[0206] NH- / and wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein surfactant 10 to surfactant 18 have the following formula:

[0207] GAR-P04610W010 Application (final). docx surfactant 10: PFPE(2000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(2000 g / mol) surfactant 11 : PFPE(4000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(4000 g / mol) surfactant 12: PFPE(7000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(7000 g / mol) surfactant 13: PFPE(2000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(2000 g / mol) surfactant 14: PFPE(4000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(4000 g / mol) surfactant 15: PFPE(7000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(7000 g / mol) surfactant 16: PFPE(2000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(2000 g / mol) surfactant 17: PFPE(4000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(4000 g / mol) surfactant 18: PFPE(7000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(7000 g / mol) wherein L2and L3represent a triazole ring having the structures: wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 95% as determined by NMR spectroscopy.

[0208] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9, wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 98% as determined by NMR spectroscopy.

[0209] Preferably, the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 96%, more preferably of at least 97%, more preferably of at least 98%, more preferably of at least 98.5%, more preferably of at least 99%, more preferably of at least 99.5%, more preferably of at least 99,6%, more preferably of at least 99.7%, more preferably of at least 99.8%, and most preferably of at least 99.9% as determined by NMR spectroscopy.

[0210] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant,

[0211] GAR-P04610W010 Application (final). docx wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9, wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7000 (1 ,1 ,1 ,2,2,3,3-heptafluoro-3-methoxy-propane), methylnonafluoro-n- butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy- butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 -ethoxy-1 ,1 ,2, 2, 3, 3,4,4, 4-nonafluorobutane and 1-ethoxy- 1 , 1 ,2,3,3,3-hexafluoro-2-(trifluoromethyl)propane), HFE-7300 (1 ,1 ,1 , 2, 2, 3, 4, 5,5,5- decafluoro-3-methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy- 1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), FC-40

[0212] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), FC-43

[0213] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(1 ,1 ,2,2,3, 3,4,4,4-nonafluorobutyl)butan-1 -amine), FC-70 (1 ,1 ,2,2,3,3,4,4,5,5,5-undecafluoro-N,N-bis(undecafluoropentyl)pentan-1-amine), FC-72 (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned, wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 98% as determined by NMR spectroscopy.

[0214] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9, wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7500 (3-ethoxy- 1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), and FC-40 (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 98% as determined by NMR spectroscopy.

[0215] GAR-P04610W010 Application (final). docx In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9, wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the total concentration of the least one amphiphilic polymer is in a range between 0.5 mM x a + 0.5 mM and 10 mM x a + 10 mM, wherein a is the molar fraction of the diblock co-polymer surfactant in the least one amphiphilic polymer, wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 98% as determined by NMR spectroscopy.

[0216] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9, wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the total concentration of the least one amphiphilic polymer is 2.5 mM x a + 2.5 mM, wherein a is the molar fraction of the diblock co-polymer surfactant in the least one amphiphilic polymer wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 98% as determined by NMR spectroscopy.

[0217] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent and two amphiphilic polymers, wherein the amphiphilic polymers are selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9, wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18,

[0218] GAR-P04610W010 Application (final). docx wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 98% as determined by NMR spectroscopy.

[0219] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant, wherein the formulation comprises three amphiphilic polymers selected from two different diblock co-polymer surfactants and a triblock co-polymer surfactant or one diblock co-polymer surfactant and two different triblock co-polymer surfactants, or wherein the formulation comprises four amphiphilic polymers selected from three different diblock co-polymer surfactants and one triblock co-polymer surfactant, or four different diblock co-polymer surfactants, or two different diblock co-polymer surfactants and two different triblock co-polymer surfactants, or one diblock co-polymer surfactant and three different triblock co-polymer surfactants, or four different triblock co-polymer surfactants, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9, wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 98% as determined by NMR spectroscopy.

[0220] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent the following diblock co-polymer surfactants and triblock co-polymer surfactants selected from, surfactant 1 surfactant 1 , surfactant 10, surfactant 11 surfactant 1 , surfactant 10, surfactant 17 surfactant 1 , surfactant 11 , surfactant 13, surfactant 15 surfactant 1 , surfactant 13, surfactant 15, surfactant 16 surfactant 1 , surfactant 2, surfactant 4, surfactant 15 surfactant 1 , surfactant 3, surfactant 7, surfactant 11 surfactant 1 , surfactant 4, surfactant 11 , surfactant 13 surfactant 1 , surfactant 4, surfactant 11 , surfactant 16 surfactant 1 , surfactant 4, surfactant 18 surfactant 1 , surfactant 6, surfactant 11 , surfactant 12 surfactant 1 , surfactant 7, surfactant 11 , surfactant 12 surfactant 1 , surfactant 7, surfactant 13, surfactant 14 surfactant 1 , surfactant 8, surfactant 15, surfactant 18 surfactant 1 , surfactant 9, surfactant 13, surfactant 16 surfactant 10 surfactant 10, surfactant 13, surfactant 14, surfactant 15

[0221] GAR-P04610W010 Application (final). docx surfactant 10, surfactant 14, surfactant 17, surfactant 18 surfactant 11 surfactant 11 , surfactant 6, surfactant 12 surfactant 11 , surfactant 9, surfactant 16 surfactant 12 surfactant 12, surfactant 13, surfactant 17 surfactant 13 surfactant 13, surfactant 14, surfactant 18 surfactant 13, surfactant 15 surfactant 13, surfactant 7, surfactant 9 surfactant 14 surfactant 14, surfactant 10, surfactant 17 surfactant 14, surfactant 3, surfactant 15 surfactant 15 surfactant 15, surfactant 12, surfactant 14 surfactant 15, surfactant 14, surfactant 18 surfactant 16 surfactant 16, surfactant 14 surfactant 16, surfactant 8, surfactant 18 surfactant 17 surfactant 17, surfactant 5, surfactant 13 surfactant 17, surfactant 8, surfactant 18 surfactant 17, surfactant 8, surfactant 9 surfactant 18 surfactant 18, surfactant 1 , surfactant 13 surfactant 18, surfactant 10, surfactant 13 surfactant 18, surfactant 14, surfactant 16 surfactant 18, surfactant 5, surfactant 11 surfactant 18, surfactant 5, surfactant 12 surfactant 18, surfactant 9, surfactant 13 surfactant 2, surfactant 11 , surfactant 18 surfactant 2, surfactant 3, surfactant 4, surfactant 10 surfactant 2, surfactant 3, surfactant 4, surfactant 12 surfactant 2, surfactant 3, surfactant 8 surfactant 2, surfactant 4, surfactant 5, surfactant 12 surfactant 2, surfactant 4, surfactant 8, surfactant 12 surfactant 2, surfactant 5, surfactant 18 surfactant 2, surfactant 6, surfactant 15, surfactant 16 surfactant 3, surfactant 1 , surfactant 15 surfactant 3, surfactant 10, surfactant 11 surfactant 3, surfactant 10, surfactant 12, surfactant 16 surfactant 3, surfactant 15, surfactant 16 surfactant 3, surfactant 2, surfactant 7 surfactant 3, surfactant 4, surfactant 10, surfactant 13 surfactant 3, surfactant 4, surfactant 16 surfactant 3, surfactant 4, surfactant 5, surfactant 16 surfactant 3, surfactant 4, surfactant 8 surfactant 3, surfactant 7, surfactant 15 surfactant 3, surfactant 8, surfactant 14, surfactant 16 surfactant 4, surfactant 5, surfactant 6 surfactant 5, surfactant 12, surfactant 16 surfactant 5, surfactant 13, surfactant 17

[0222] GAR-P04610W010 Application (final). docx surfactant 5, surfactant 14 surfactant 5, surfactant 6, surfactant 11 , surfactant 15 surfactant 5, surfactant 7, surfactant 12, surfactant 14 surfactant 6 surfactant 6, surfactant 13, surfactant 17 surfactant 6, surfactant 15 surfactant 7 surfactant 7, surfactant 1 surfactant 7, surfactant 1 , surfactant 10 surfactant 7, surfactant 10, surfactant 17 surfactant 7, surfactant 11 , surfactant 12, surfactant 16 surfactant 7, surfactant 12, surfactant 13, surfactant 17 surfactant 7, surfactant 13, surfactant 14, surfactant 17 surfactant 7, surfactant 16 surfactant 7, surfactant 8, surfactant 13, surfactant 15 surfactant 8 surfactant 8, surfactant 1 , surfactant 18 surfactant 8, surfactant 10, surfactant 11 , surfactant 12 surfactant 8, surfactant 11 , surfactant 14, surfactant 15 surfactant 8, surfactant 12, surfactant 14, surfactant 15 surfactant 8, surfactant 17 surfactant 8, surfactant 3, surfactant 15 surfactant 8, surfactant 9, surfactant 10, surfactant 14 surfactant 9 surfactant 9, surfactant 11 , surfactant 12, surfactant 16 surfactant 9, surfactant 17 surfactant 9, surfactant 18 surfactant 9, surfactant 2, surfactant 4. wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 98% as determined by NMR spectroscopy. In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent the following diblock co-polymer surfactants and triblock co-polymer surfactants selected from surfactants molar ratio surfactant 1 surfactant 1 , surfactant 10, surfactant 11 2:1 :1 surfactant 1 , surfactant 10, surfactant 17 2:1 :1 surfactant 1 , surfactant 2, surfactant 4, surfactant 15 1 : 1 : 1 : 1 surfactant 1 , surfactant 3, surfactant 7, surfactant 11 1 : 1 : 1 : 1 surfactant 1 , surfactant 4, surfactant 11 , surfactant 13 1 : 1 : 1 : 1 surfactant 1 , surfactant 4, surfactant 11 , surfactant 16 1 : 1 : 1 : 1 surfactant 1 , surfactant 4, surfactant 18 2:1 :1 surfactant 1 , surfactant 6, surfactant 11 , surfactant 12 1 : 1 : 1 : 1 surfactant 1 , surfactant 7, surfactant 11 , surfactant 12 1 : 1 : 1 : 1

[0223] GAR-P04610W010 Application (final). docx ABK 43 surfactant 1 , surfactant 7, surfactant 13, surfactant 14 1 : 1 : 1 : 1 surfactant 1 , surfactant 8, surfactant 15, surfactant 18 1 : 1 : 1 : 1 surfactant 1 , surfactant 9, surfactant 13, surfactant 16 1 : 1 : 1 : 1 surfactant 11 , surfactant 6, surfactant 12 2:1 :1 surfactant 13 surfactant 13, surfactant 7, surfactant 9 2:1 :1 surfactant 14, surfactant 3, surfactant 15 2:1 :1 surfactant 17, surfactant 5, surfactant 13 2:1 :1 surfactant 17, surfactant 8, surfactant 9 2:1 :1 surfactant 18, surfactant 5, surfactant 11 2:1 :1 surfactant 18, surfactant 5, surfactant 12 2:1 :1 surfactant 2, surfactant 11 , surfactant 18 2:1 :1 surfactant 2, surfactant 3, surfactant 4, surfactant 10 1 : 1 : 1 : 1 surfactant 2, surfactant 3, surfactant 4, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 3, surfactant 8 2:1 :1 surfactant 2, surfactant 4, surfactant 5, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 4, surfactant 8, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 5, surfactant 18 2:1 :1 surfactant 2, surfactant 6, surfactant 15, surfactant 16 1 : 1 : 1 : 1 surfactant 3, surfactant 1 , surfactant 15 2:1 :1 surfactant 3, surfactant 4, surfactant 10, surfactant 13 1 : 1 : 1 : 1 surfactant 3, surfactant 4, surfactant 5, surfactant 16 1 : 1 : 1 : 1 surfactant 3, surfactant 4, surfactant 8 2:1 :1 surfactant 3, surfactant 7, surfactant 15 2:1 :1 surfactant 3, surfactant 8, surfactant 14, surfactant 16 1 : 1 : 1 : 1 surfactant 4, surfactant 5, surfactant 6 2:1 :1 surfactant 5, surfactant 12, surfactant 16 2:1 :1 surfactant 5, surfactant 13, surfactant 17 2:1 :1 surfactant 5, surfactant 14 1 :1 surfactant 5, surfactant 6, surfactant 11 , surfactant 15 1 : 1 : 1 : 1 surfactant 5, surfactant 7, surfactant 12, surfactant 14 1 : 1 : 1 : 1 surfactant 6 surfactant 6, surfactant 13, surfactant 17 2:1 :1 surfactant 6, surfactant 15 1 :1 surfactant 7 surfactant 7, surfactant 1 3:1 surfactant 7, surfactant 1 , surfactant 10 2:1 :1 surfactant 7, surfactant 10, surfactant 17 2:1 :1 surfactant 7, surfactant 13, surfactant 14, surfactant 17 1 : 1 : 1 : 1 surfactant 7, surfactant 8, surfactant 13, surfactant 15 1 : 1 : 1 : 1

[0224] GAR-P04610W010 Application (final). docx surfactant 8 surfactant 8, surfactant 1 , surfactant 18 2:1 :1 surfactant 8, surfactant 17 1 :1 surfactant 8, surfactant 10, surfactant 11 , surfactant 12 1 : 1 : 1 : 1 surfactant 8, surfactant 11 , surfactant 14, surfactant 15 1 : 1 : 1 : 1 surfactant 8, surfactant 12, surfactant 14, surfactant 15 1 : 1 : 1 : 1 surfactant 8, surfactant 9, surfactant 10, surfactant 14 1 : 1 : 1 : 1 surfactant 9, surfactant 2, surfactant 4 2:1 :1 wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 98% as determined by NMR spectroscopy.

[0225] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent the following diblock co-polymer surfactants and triblock co-polymer surfactants selected from, surfactants molar ratio surfactant 1 surfactant 2, surfactant 4, surfactant 5, surfactant 12 1 :1 :1 :1 surfactant 2, surfactant 4, surfactant 8, surfactant 12 1 :1 :1 :1 surfactant 4, surfactant 5, surfactant 6 2:1 :1. wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 98% as determined by NMR spectroscopy.

[0226] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9, wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the formulation is devoid of free perfluoropolyether carboxylic acid, wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 98% as determined by NMR spectroscopy.

[0227] In a preferred embodiment, the formulation comprises a water-immiscible fluorocarbon solvent, at least one amphiphilic polymer further comprising 0.1 mM to 15 mM of a

[0228] GAR-P04610W010 Application (final). docx perfluoropolyether carboxylic acid, wherein the amphiphilic polymer is selected from a diblock co-polymer surfactant and a triblock co-polymer surfactant, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9, wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein the diblock co-polymer surfactant and the triblock co-polymer surfactant have a purity of at least 98% as determined by NMR spectroscopy.

[0229] Rather than employing a singular surfactant, the present invention utilizes a comprehensive array of surfactant formulations of distinct surfactants. These surfactants were characterized by variations in the molecular weights of their polyethylene glycol (PEG) and perfluoropolyether (PFPE) segments, in addition to the synthesis of both diblock and triblock copolymers. Each surfactant exhibited unique characteristics, notably in aspects such as interfacial tension (IFT) reduction, velocity of surfactant layer formation, and droplet stabilization capabilities. This diverse collection enables the formulation of multi-surfactant mixtures, tailored to optimize specific properties to meet experimental requirements.

[0230] Among these distinct surfactant molecules, they can be amalgamated in various ratios comprising two or more molecules into surfactant formulations for three primary applications within droplet-based microfluidics: (1 ) construction of synthetic cells, (2) encapsulation of cells, and (3) droplet digital molecular biology.

[0231] (1) Synthetic Cell Construction:

[0232] This approach involves the encapsulation of materials for synthetic cell formation within droplets. A critical aspect of this method is the addition of Krytox at precise concentrations to facilitate a charge-induced self-assembly of lipid bilayers at the wateroil interface. Insufficient Krytox concentrations fail to form a lipid layer, whereas excessive concentrations detrimentally affect the encapsulated materials, such as proteins, by lowering the pH. Furthermore, post-incubation and lipid bilayer formation, the synthetic cells must be transitioned from an oil / water to a water / water system, necessitating a surfactant that forms a metastable system — sufficiently robust for stabilization yet amenable to destabilization for cell release.

[0233] Thus, another aspect of the present invention is directed to a formulation for the synthetic cell formation within droplets comprising a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer

[0234] GAR-P04610W010 Application (final). docx is selected from a diblock co-polymer surfactant of formula A-B and a triblock copolymer surfactant of formula B’-A’-B’, wherein A and A’ independently of each other represent a water-soluble polymer block having a range in molecular weight from 300 g / mol to 2000 g / mol, and B and B’ represent independently of each other a perfluorinated polyether block having a range in molecular weight from 1500 g / mol to 7500 g / mol.

[0235] (2) Cell Encapsulation:

[0236] This process encapsulates cells along with cell media and nutrients, such as fetal bovine serum (FBS), creating a high osmotic pressure that enhances droplet fusion forces. The encapsulated cells are also incubated at 37 °C, imposing additional requirements on the surfactant formulation for stability under these conditions. Moreover, a Krytox-free environment is essential due to its cytotoxicity.

[0237] Thus, another aspect of the present invention is directed to a formulation for the cell encapsulation and cell stabilization comprising a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant of formula A-B and a triblock co-polymer surfactant of formula B’-A’-B’, wherein A and A’ independently of each other represent a water-soluble polymer block having a range in molecular weight from 300 g / mol to 2000 g / mol, and B and B’ represent independently of each other a perfluorinated polyether block having a range in molecular weight from 1500 g / mol to 7500 g / mol.

[0238] (3) Droplet Digital Molecular Biology:

[0239] For conducting chemical reactions and analyzing biomolecules within droplets, as in droplet digital PCR analysis, surfactants must ensure droplet stability under stringent chemical reaction conditions, including elevated temperatures. These analyses are quantitative, and droplet fusion impacts the accuracy of such analyses. During the material analysis process, droplets undergo physical manipulation through pipetting or reinjection, subjecting them to considerable shear stress, which they must withstand.

[0240] Thus, another aspect of the present invention is directed to a formulation for analyzing biomolecules within droplets comprising a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant of formula A-B and a triblock co-polymer surfactant of formula B’-A’-B’, wherein A and A’ independently of each other represent a water- soluble polymer block having a range in molecular weight from 300 g / mol to 2000 g / mol, and B and B’ represent independently of each other a perfluorinated polyether block having a range in molecular weight from 1500 g / mol to 7500 g / mol.

[0241] GAR-P04610W010 Application (final). docx Fig. 1.1 depicts a schematic representation of a diblock copolymer surfactant, where the water-soluble polymer block is shown as a dashed line and the perfluorinated polymer block as a solid line. This list of polymers could be used as water-soluble polymer block, but not limited to: polyethylene glycol, polyvinyl alcohol, polyacrylic acid, polymethacrylic acid, polyvinylpyrrolidone, carboxymethyl cellulose, hydroxyethyl cellulose, and polyacrylamide. Perfluorinated polyether could be used as perfluorinated polymer block, but not limited to it. Fig. 1.2 depicts a schematic representation of a triblock copolymer surfactant, where the water-soluble polymer block is shown as a dashed line and the perfluorinated polymer block as a solid line. This list of polymers could be used as water-soluble polymer block, but not limited to: polyethylene glycol, polyvinyl alcohol, polyacrylic acid, polymethacrylic acid, polyvinylpyrrolidone, carboxymethyl cellulose, hydroxyethyl cellulose, and polyacrylamide. Perfluorinated polyether could be used as perfluorinated polymer block, but not limited to it. Fig. 1.3 depicts the molecular structure of perfluorinated polyether as an example for the perfluorinated polymer blocks. The dashed line represents the connection the rest of the block copolymer. Fig. 1.4 depicts the molecular structure of polyethylene glycol as an example for the water- soluble polymer blocks. The dashed lines represent possible connections the rest of the block copolymer. Fig. 1 .5 to Fig. 1 .9 depicts a list of functional groups, which could be used to link the blocks of the block copolymers, but not limited to: ether group (Fig. 1 .5), ester group (Fig. 1.6), amine group (Fig. 1.7), amide group (Fig. 1.8), or triazole ring (Fig. 1.9).

[0242] Fig. 2.1 depicts a comparative measurement of the interfacial tension between water and fluorinated oil, in this case 3MTMNovec™ 7500, with dissolved block copolymer surfactant. The blue line shows the automatic fit, which is the basis for calculating the interfacial tension using the Young-Laplace model. Fig. 2.2 depicts an exemplary plot of the time-depended change of interfacial tension. The x-axis represents the time in minutes after the hanging drop was created. The y-axis represents the calculated interfacial tension in milli Newton per meter. The time-dependent interfacial tension was measured three times. The solid line represents the mean value and the area with reduced opacity the standard deviation.

[0243] Fig. 3.1 - Fig. 3.6 depicts microscopy images of w / o droplets, generated with two different surfactant formulations. The droplets were imaged immediately after production, after 24 hours, and after 48 hours. Fig. 3.1 - Fig. 3.3 depicts the analysis where a surfactant formulation consisting of 60 % diblock copolymer with PEG with a molecular weight of 550 g / mol as water-soluble polymer block and PFPE with a molecular weight of 4000 g / mol as perfluorinated polymer block, and 40 % diblock copolymer with PEG with a molecular weight of 550 g / mol as water-soluble polymer block and PFPE with a molecular weight of 7000 g / mol as perfluorinated polymer block

[0244] GAR-P04610W010 Application (final). docx was used. The polydispersity index of the shown w / o droplets is low and is not increasing significantly over time. Fig. 3.4 - Fig. 3.6 depicts the analysis where a surfactant formulation consisting of 100 % triblock copolymer with PEG with a molecular weight of 600 g / mol as water-soluble polymer block and PFPE with a molecular weight of 4000 g / mol as perfluorinated polymer block was used. The polydispersity index of the shown w / o droplets is increasing overtime since the w / o droplets are fusing.

[0245] Fig. 4 depicts a comparative plot of the lipid vesicle release yield from w / o droplet stabilized lipid vesicles. The x-axis represents the different conditions under which the lipid vesicle release yield was determined. The y-axis represents the measured phospholipid concentration correlating with the amount of lipid vesicles. The measurement of the concentration of the small unilamellar vesicles (SUV) shows the maximal possible concentration since this lipid concentration was used to create the giant unilamellar vesicles (GUV). Using the exemplary formulation 1 , the amount of lipid vesicles after release was higher than comparing with using the exemplary formulation 2 conducting that formulation 1 is a better formulation for constructing lipid vesicles.

[0246] Fig. 5 depicts a comparative violin plot showcasing the viability of cells encapsulated within w / o droplets, monitored over various incubation periods. The x-axis quantifies the incubation duration post-encapsulation, while the y-axis displays the normalized values of cell viability, benchmarked against the immediate post-encapsulation readings. Notably, the trajectory associated with Formulation 1 exhibits a slower decline in viability over time in comparison to Formulation 2. This indicates a superior preservation of cell viability under Formulation 1 , thereby extending the feasible window for conducting detailed observations in single-cell experiments.

[0247] Fig. 6 depicts a comparative scatter plot representing the fluorescent readout of a molecular biology essay executed in bulk or in w / o droplets using different surfactant formulations. This illustration is an example of various biological detection methods based on the measurement of fluorescence signals, such as droplet digital PCR. The x- axis represents the different conditions under which the experiment was performed, while the y-axis shows the fluorescence values of the experiment in correlation with the amount of target molecules. The Bulk condition was performed in a 96-well plate, while the analysis under the conditions of Formulation 1 and 2 was performed in w / o drops using the corresponding formulations. Using Formulation 1 , the same value is achieved like the bulk analysis. Due to the high-throughput analysis, more data points could be analyzed, which supports the strength of the scientific interpretation. When using formulation 2, the mean fluorescence value deviates considerably from the bulk analysis, and the fluctuation of the values is also significantly greater.

[0248] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 1 .

[0249] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 4, 18.

[0250] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 10, 11.

[0251] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 10, 17.

[0252] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 2, 4, 15.

[0253] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 3, 1 , 15.

[0254] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 3, 7, 11.

[0255] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 4, 11 , 13.

[0256] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 4, 11 , 16.

[0257] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 6, 11 , 12.

[0258] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 7, 1.

[0259] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 7, 1 , 10.

[0260] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 7, 11 , 12.

[0261] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 7, 13, 14.

[0262] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 8, 1 , 18.

[0263] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 8, 15, 18.

[0264] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 9, 13, 16.

[0265] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 11 , 13, 15.

[0266] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 1 , 13, 15, 16.

[0267] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 18, 1 , 13.

[0268] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 2, 3, 8.

[0269] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 2, 5, 18.

[0270] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 2, 11 , 18.

[0271] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 3, 2, 7.

[0272] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 2, 3, 4, 10.

[0273] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 2, 3, 4, 12.

[0274] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 2, 4, 5, 12.

[0275] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 2, 4, 8, 12.

[0276] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 9, 2, 4.

[0277] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 2, 6, 15, 16.

[0278] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 3, 4, 8.

[0279] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 3, 4, 16.

[0280] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 3, 7, 15.

[0281] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 3, 10, 11.

[0282] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 3, 15, 16.

[0283] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 3, 4, 5, 16.

[0284] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 3, 4, 10, 13.

[0285] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 8, 3, 15.

[0286] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 3, 8, 14, 16.

[0287] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 3, 10, 12, 16.

[0288] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 14, 3, 15.

[0289] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 4, 5, 6.

[0290] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 5, 12, 16.

[0291] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 5, 13, 17.

[0292] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 5, 14.

[0293] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 5, 6, 11 , 15.

[0294] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 5, 7, 12, 14.

[0295] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 18, 5, 11.

[0296] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 18, 5, 12.

[0297] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 17, 5, 13.

[0298] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 6.

[0299] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 6, 13, 17.

[0300] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 6, 15.

[0301] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 11 , 6, 12.

[0302] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 7.

[0303] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 7, 10, 17.

[0304] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 7, 16.

[0305] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 7, 8, 13, 15.

[0306] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 13, 7, 9.

[0307] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 7, 11 , 12, 16.

[0308] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 7, 12, 13, 17.

[0309] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 7, 13, 14, 17.

[0310] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 8.

[0311] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 8, 17.

[0312] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 8, 9, 10, 14.

[0313] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 17, 8, 9.

[0314] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 8, 10, 11 , 12.

[0315] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 8, 11 , 14, 15.

[0316] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 8, 12, 14, 15.

[0317] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 16, 8, 18.

[0318] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 17, 8, 18.

[0319] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 9.

[0320] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 9, 17.

[0321] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 9, 18.

[0322] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 11 , 9, 16.

[0323] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 9, 11 , 12, 16.

[0324] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 18, 9, 13.

[0325] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 10.

[0326] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 10, 13, 14, 15.

[0327] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 18, 10, 13.

[0328] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 14, 10, 17.

[0329] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 10, 14, 17, 18.

[0330] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 11.

[0331] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 12.

[0332] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 12, 13, 17.

[0333] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 15, 12, 14.

[0334] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 13.

[0335] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 13, 14, 18.

[0336] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 13, 15.

[0337] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 14.

[0338] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 15, 14, 18.

[0339] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 16, 14.

[0340] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymers surfactants 18, 14, 16.

[0341] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 15.

[0342] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 16.

[0343] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 17.

[0344] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer surfactant 18.

[0345] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 4, surfactant 18 in a molar ratio of 2:1 :1.

[0346] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 10, surfactant 11 in a molar ratio of 2:1 :1.

[0347] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 10, surfactant 17 in a molar ratio of 2: 1 : 1 .

[0348] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 2, surfactant 4, surfactant 15 in a molar ratio of 1 : 1 : 1 : 1 .

[0349] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 3, surfactant 1 , surfactant 15 in a molar ratio of 2:1 :1.

[0350] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 3, surfactant 7, surfactant 11 in a molar ratio of 1 : 1 : 1 : 1 .

[0351] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 4, surfactant 11 , surfactant 13 in a molar ratio of 1 : 1 : 1 : 1 .

[0352] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 4, surfactant 11 , surfactant 16 in a molar ratio of 1 : 1 : 1 : 1 .

[0353] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 6, surfactant 11 , surfactant 12 in a molar ratio of 1 : 1 : 1 : 1 .

[0354] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 7, surfactant 1 in a molar ratio of 3:1 .

[0355] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 7, surfactant 1 , surfactant 10 in a molar ratio of 2:1 :1.

[0356] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 7, surfactant 11 , surfactant 12 in a molar ratio of 1 : 1 : 1 : 1 .

[0357] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 7, surfactant 13, surfactant 14 in a molar ratio of 1 : 1 : 1 : 1 .

[0358] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 8, surfactant 1 , surfactant 18 in a molar ratio of 2:1 :1.

[0359] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 8, surfactant 15, surfactant 18 in a molar ratio of 1 : 1 : 1 : 1 .

[0360] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 9, surfactant 13, surfactant 16 in a molar ratio of 1 : 1 : 1 : 1 .

[0361] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 11 , surfactant 13, surfactant 15 in a molar ratio of 1 : 1 : 1 : 1 .

[0362] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 1 , surfactant 13, surfactant 15, surfactant 16 in a molar ratio of 1 : 1 : 1 : 1 .

[0363] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 18, surfactant 1 , surfactant 13 in a molar ratio of 2: 1 : 1 .

[0364] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 2, surfactant 3, surfactant 8 in a molar ratio of 2:1 :1.

[0365] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 2, surfactant 5, surfactant 18 in a molar ratio of 2:1 :1.

[0366] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 2, surfactant 11 , surfactant 18 in a molar ratio of 2: 1 : 1 .

[0367] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 3, surfactant 2, surfactant 7 in a molar ratio of 2:1 :1.

[0368] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 2, surfactant 3, surfactant 4, surfactant 10 in a molar ratio of 1 : 1 : 1 : 1 .

[0369] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 2, surfactant 3, surfactant 4, surfactant 12 in a molar ratio of 1 : 1 : 1 : 1 .

[0370] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 2, surfactant 4, surfactant 5, surfactant 12 in a molar ratio of 1 : 1 : 1 : 1 .

[0371] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 2, surfactant 4, surfactant 8, surfactant 12 in a molar ratio of 1 : 1 : 1 : 1 .

[0372] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 9, surfactant 2, surfactant 4 in a molar ratio of 2:1 :1.

[0373] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 2, surfactant 6, surfactant 15, surfactant 16 in a molar ratio of 1 : 1 : 1 : 1 .

[0374] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 3, surfactant 4, surfactant 8 in a molar ratio of 2:1 :1.

[0375] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 3, surfactant 4, surfactant 16 in a molar ratio of 2:1 :1.

[0376] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 3, surfactant 7, surfactant 15 in a molar ratio of 2:1 :1.

[0377] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 3, surfactant 10, surfactant 11 in a molar ratio of 2:1 :1.

[0378] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 3, surfactant 15, surfactant 16 in a molar ratio of 2: 1 : 1 .

[0379] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 3, surfactant 4, surfactant 5, surfactant 16 in a molar ratio of 1 : 1 : 1 : 1 .

[0380] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 3, surfactant 4, surfactant 10, surfactant 13 in a molar ratio of 1 : 1 : 1 : 1 .

[0381] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 8, surfactant 3, surfactant 15 in a molar ratio of 2:1 :1.

[0382] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 3, surfactant 8, surfactant 14, surfactant 16 in a molar ratio of 1 : 1 : 1 : 1 .

[0383] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 3, surfactant 10, surfactant 12, surfactant 16 in a molar ratio of 1 : 1 : 1 : 1 .

[0384] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 14, surfactant 3, surfactant 15 in a molar ratio of 2: 1 : 1 .

[0385] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 4, surfactant 5, surfactant 6 in a molar ratio of 2:1 :1.

[0386] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 5, surfactant 12, surfactant

[0387] 16 in a molar ratio of 2: 1 : 1 .

[0388] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 5, surfactant 13, surfactant

[0389] 17 in a molar ratio of 2: 1 : 1 .

[0390] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 5, surfactant 14 in a molar ratio of 1 : 1 .

[0391] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 5, surfactant 6, surfactant

[0392] 11 , surfactant 15 in a molar ratio of 1 : 1 : 1 : 1 .

[0393] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 5, surfactant 7, surfactant

[0394] 12, surfactant 14 in a molar ratio of 1 : 1 : 1 : 1 .

[0395] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 18, surfactant 5, surfactant

[0396] 11 in a molar ratio of 2:1 :1.

[0397] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 18, surfactant 5, surfactant

[0398] 12 in a molar ratio of 2: 1 : 1 .

[0399] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 17, surfactant 5, surfactant

[0400] 13 in a molar ratio of 2: 1 : 1 .

[0401] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 6, surfactant 13, surfactant 17 in a molar ratio of 2: 1 : 1 .

[0402] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 6, surfactant 15 in a molar ratio of 1 : 1 .

[0403] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 11 , surfactant 6, surfactant 12 in a molar ratio of 2: 1 : 1 .

[0404] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 7, surfactant 10, surfactant 17 in a molar ratio of 2: 1 : 1 .

[0405] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 7, surfactant 16 in a molar ratio of 1 : 1 .

[0406] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 7, surfactant 8, surfactant 13, surfactant 15 in a molar ratio of 1 : 1 : 1 : 1 .

[0407] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 13, surfactant 7, surfactant 9 in a molar ratio of 2:1 :1.

[0408] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 7, surfactant 11 , surfactant

[0409] 12, surfactant 16 in a molar ratio of 1 : 1 : 1 : 1 .

[0410] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 7, surfactant 12, surfactant

[0411] 13, surfactant 17 in a molar ratio of 1 : 1 : 1 : 1 .

[0412] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 7, surfactant 13, surfactant

[0413] 14, surfactant 17 in a molar ratio of 1 : 1 : 1 : 1 .

[0414] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 8, surfactant 17 in a molar ratio of 1 : 1 .

[0415] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 8, surfactant 9, surfactant

[0416] 10, surfactant 14 in a molar ratio of 1 : 1 : 1 : 1 .

[0417] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 17, surfactant 8, surfactant 9 in a molar ratio of 2:1 :1.

[0418] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 8, surfactant 10, surfactant

[0419] 11 , surfactant 12 in a molar ratio of 1 : 1 : 1 : 1 .

[0420] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 8, surfactant 11 , surfactant 14, surfactant 15 in a molar ratio of 1 : 1 : 1 : 1 .

[0421] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 8, surfactant 12, surfactant 14, surfactant 15 in a molar ratio of 1 : 1 : 1 : 1 .

[0422] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 16, surfactant 8, surfactant 18 in a molar ratio of 2: 1 : 1 .

[0423] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 17, surfactant 8, surfactant 18 in a molar ratio of 2: 1 : 1 .

[0424] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 9, surfactant 17 in a molar ratio of 1 : 1 .

[0425] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 9, surfactant 18 in a molar ratio of 1 : 1 .

[0426] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 11 , surfactant 9, surfactant

[0427] 16 in a molar ratio of 2: 1 : 1 .

[0428] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 9, surfactant 11 , surfactant 12, surfactant 16 in a molar ratio of 1 : 1 : 1 : 1 .

[0429] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 18, surfactant 9, surfactant 13 in a molar ratio of 2: 1 : 1 .

[0430] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 10, surfactant 13, surfactant 14, surfactant 15 in a molar ratio of 1 : 1 : 1 : 1 .

[0431] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 18, surfactant 10, surfactant 13 in a molar ratio of 2: 1 : 1 .

[0432] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 14, surfactant 10, surfactant

[0433] 17 in a molar ratio of 2: 1 : 1 .

[0434] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 10, surfactant 14, surfactant 17, surfactant 18 in a molar ratio of 1 : 1 : 1 : 1 .

[0435] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 12, surfactant 13, surfactant

[0436] 17 in a molar ratio of 2: 1 : 1 .

[0437] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 15, surfactant 12, surfactant in a molar ratio of 2: 1 : 1 .

[0438] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 13, surfactant 14, surfactant

[0439] 18 in a molar ratio of 2: 1 : 1 .

[0440] GAR-P04610W010 Application (final). docx In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 13, surfactant 15 in a molar ratio of 1 : 1 .

[0441] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 15, surfactant 14, surfactant 18 in a molar ratio of 2: 1 : 1 .

[0442] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 16, surfactant 14 in a molar ratio of 3:1 .

[0443] In a preferred embodiment, the inventive formulation comprises a water-immiscible fluorocarbon solvent and amphiphilic polymer(s) surfactant 18, surfactant 14, surfactant 16 in a molar ratio of 2: 1 : 1 .

[0444] Preferably, in all aforementioned embodiments, the total concentration of the amphiphilic polymer is in the range between 1 mM and 20 mM, preferably between 4 mM and 6 mM and most preferably 5 mM, when the amphiphilic polymer is selected from surfactant 1 to 9. Preferably, the total concentration of the amphiphilic polymer is in the range between 0.5 mM and 10 mM, preferably between 1.5 mM and 3.5 mM and most preferably 2.5 mM, when the amphiphilic polymer is selected from surfactant 10 to 18.

[0445] Preferably, in all aforementioned embodiments, the concentration of each amphiphilic polymer is in the range between 1 mM and 20 mM, preferably between 4 mM and 6 mM and most preferably 5 mM, when the amphiphilic polymer is selected from surfactant 1 to 9. Preferably, the concentration of each amphiphilic polymer is in the range between 0.5 mM and 10 mM, preferably between 1.5 mM and 3.5 mM and most preferably 2.5 mM, when the amphiphilic polymer is selected from surfactant 10 to 18.

[0446] Preferably, in all aforementioned embodiments, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7000 (1 , 1 , 1 , 2,2,3, 3-heptafluoro-3- methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2- (trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 -ethoxy-1 , 1 ,2, 2, 3, 3, 4,4,4- nonafluorobutane and 1 -ethoxy-1 , 1 ,2, 3, 3, 3-hexafluoro-2-(trifluoromethyl)propane), HFE- 7300 (1 ,1 ,1 ,2,2,3,4,5,5,5-decafluoro-3-methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), FC-40

[0447] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), FC-43

[0448] (1 ,1 ,2,2,3, 3,4,4,4-nonafluoro-N,N-bis(1 ,1 ,2,2,3, 3,4,4,4-nonafluorobutyl)butan-1 -amine), FC-70 (1 ,1 ,2,2,3,3,4,4,5,5,5-undecafluoro-N,N-bis(undecafluoropentyl)pentan-1-amine),

[0449] GAR-P04610W010 Application (final). docx FC-72 (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned.

[0450] More preferably, in all aforementioned embodiments, the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 ,1 ,1 ,2,3,3-hexafluoro-3-methoxy-2- (trifluoromethyl)propane), HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2- (trifluoromethyl)hexane), and FC-40 (1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N,N- bis(nonafluorobutyl)butan-1 -amine).

[0451] Preferably, in all aforementioned embodiments, the water-immiscible fluorocarbon solvent is HFE-7500 (3-ethoxy-1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2- (trifluoromethyl)hexane).

[0452] Preferably, in all aforementioned embodiments, the inventive formulation is devoid of free perfluoropolyether carboxylic acid.

[0453] Preferably, in all aforementioned embodiments, the formulation further comprises 0.1 mM to 15 mM of a perfluoropolyether carboxylic acid. Preferably, the perfluoropolyether carboxylic acid is a polymer of hexafluoropropylene oxide.

[0454] Preferably, in all aforementioned embodiments, the diblock co-polymer surfactants and the triblock co-polymer surfactants have a purity of at least 95% as determined by NMR spectroscopy.

[0455] Preferably, in all aforementioned embodiments, the diblock co-polymer surfactants and the triblock co-polymer surfactants have a purity of at least 96%, more preferably of at least 97%, more preferably of at least 98%, more preferably of at least 98.5%, more preferably of at least 99%, more preferably of at least 99.5%, more preferably of at least 99,6%, more preferably of at least 99.7%, more preferably of at least 99.8%, and most preferably of at least 99.9% as determined by NMR spectroscopy.

[0456] Description of the Figures

[0457] Figure 1 is a schematic representation and molecular structure of diblock copolymer surfactants and triblock copolymer surfactants (water-soluble polymer block is shown as a dashed line and the perfluorinated polymer block as a solid line): 1 ) schematic representation of a diblock co-polymer; 2) schematic representation of a triblock co-

[0458] GAR-P04610W010 Application (final). docx polymer; 3) shows molecular structure of perfluorinated polyether as an example for the perfluorinated polymer blocks. The dashed line represents the connection the rest of the block copolymer; 4) shows the molecular structure of polyethylene glycol as an example for the water-soluble polymer blocks. The dashed lines represent possible connections the rest of the block copolymer; 5) - 9) possible functional groups connecting the blocks in the co-polymer: 5) ether group, 6) ester group, 7) amine group, 8) amide group, or 9) triazole ring.

[0459] Figure 2 is a representation of the pendent drop experiment and how the interfacial tension is calculated. 1 ) shows a comparative measurement of the interfacial tension between water and fluorinated oil, in this case 3M Novec 7500, with dissolved block copolymer surfactant. The blue line shows the automatic fit, which is the basis for calculating the interfacial tension using the Young-Laplace model. 2) shows an exemplary plot of the time-depended change of interfacial tension. The x-axis represents the time in minutes after the hanging drop was created. The y-axis represents the calculated interfacial tension in milli Newton per meter. The timedependent interfacial tension was measured three times. The solid line represents the mean value and the area with reduced opacity the standard deviation.

[0460] Figure 3 is exemplary images of w / o droplet produced with two different surfactants at different time points to analyze the fusion rate. 1 ) - 6) show microscopy images of w / o droplets, generated with two different surfactant formulations. The droplets were imaged immediately after production, after 24 hours, and after 48 hours. 1 ) -3) depicts the analysis where a surfactant formulation consisting of 60 % diblock copolymer with PEG with a molecular weight of 550 g / mol as water-soluble polymer block and PFPE with a molecular weight of 4000 g / mol as perfluorinated polymer block, and 40 % diblock copolymer with PEG with a molecular weight of 550 g / mol as water-soluble polymer block and PFPE with a molecular weight of 7000 g / mol as perfluorinated polymer block was used. The polydispersity index of the shown w / o droplets is low and is not increasing significantly over time. 4) - 6) depicts the analysis where a surfactant formulation consisting of 100 % triblock copolymer with PEG with a molecular weight of 600 g / mol as water-soluble polymer block and PFPE with a molecular weight of 4000 g / mol as perfluorinated polymer block was used. The polydispersity index of the shown w / o droplets is increasing overtime since the w / o droplets are fusing.

[0461] Figure 4 is a simulated violine plot representing the release efficiency difference by using different surfactant formulations. Figure 4 depicts a comparative plot of the lipid vesicle release yield from w / o droplet stabilized lipid vesicles. The x-axis represents the different conditions under which the lipid vesicle release yield was determined. The y- axis represents the measured phospholipid concentration correlating with the amount of

[0462] GAR-P04610W010 Application (final). docx lipid vesicles. The measurement of the concentration of the small unilamellar vesicles (SUV) shows the maximal possible concentration since this lipid concentration was used to create the giant unilamellar vesicles (GUV). Using the exemplary formulation 1 , the amount of lipid vesicles after release was higher than comparing with using the exemplary formulation 2 conducting that formulation 1 is a better formulation for constructing lipid vesicles.

[0463] Figure 5 is a simulated violine plot representing the difference in viability of encapsulated cells by using different surfactant formulations. Figure 5 depicts a comparative violin plot showcasing the viability of cells encapsulated within w / o droplets, monitored over various incubation periods. The x-axis quantifies the incubation duration post-encapsulation, while the y-axis displays the normalized values of cell viability, benchmarked against the immediate post-encapsulation readings. Notably, the trajectory associated with Formulation 1 exhibits a slower decline in viability over time in comparison to Formulation 2. This indicates a superior preservation of cell viability under Formulation 1 , thereby extending the feasible window for conducting detailed observations in single-cell experiments.

[0464] Figure 6 is a simulated scatter plot representing the fluorescent readout of a molecular biology assay executed in bulk or in w / o droplets using different surfactant formulations. Figure 6 depicts a comparative scatter plot representing the fluorescent readout of a molecular biology essay executed in bulk or in w / o droplets using different surfactant formulations. This illustration is an example of various biological detection methods based on the measurement of fluorescence signals, such as droplet digital PCR. The x- axis represents the different conditions under which the experiment was performed, while the y-axis shows the fluorescence values of the experiment in correlation with the amount of target molecules. The Bulk condition was performed in a 96-well plate, while the analysis under the conditions of Formulation 1 and 2 was performed in w / o drops using the corresponding formulations. Using Formulation 1 , the same value is achieved like the bulk analysis. Due to the high-throughput analysis, more data points could be analyzed, which supports the strength of the scientific interpretation. When using formulation 2, the mean fluorescence value deviates considerably from the bulk analysis, and the fluctuation of the values is also significantly greater.

[0465] The following examples are included to demonstrate preferred embodiments of the invention. It should be appreciated by those of skill in the art that the techniques disclosed in the following examples represent techniques discovered by the inventor to function well in the practice of the invention, and thus can be considered to constitute preferred modes for its practice. However, those of skill in the art should, in light of the present disclosure, appreciate that many changes can be made in the specific

[0466] GAR-P04610W010 Application (final). docx embodiments which are disclosed and still obtain a like or similar result without departing from the scope of the invention.

[0467] Further modifications and alternative embodiments of various aspects of the invention will be apparent to those skilled in the art in view of this description. Accordingly, this description is to be construed as illustrative only and is for the purpose of teaching those skilled in the art the general manner of carrying out the invention. It is to be understood that the forms of the invention shown and described herein are to be taken as examples of embodiments. Elements and materials may be substituted for those illustrated and described herein, parts and processes may be reversed, and certain features of the invention may be utilized independently, all as would be apparent to one skilled in the art after having the benefit of this description of the invention. Changes may be made in the elements described herein without departing from the scope of the invention as described in the following claims.

[0468] Examples

[0469] Example 1 : Synthesis of the block co-polymers

[0470] The diblock and triblock copolymers were synthesized according to standard routines described in the literature. For the convenience of the reader and to ensure accessibility, the fundamental steps of this synthesis are summarized here on the example of the triblock copolymer with the structure B-A’-B, where A’ is PEG with a molecular size of 1500 g / mol and B is PFPE with a molecular size of 7000 g / mol.

[0471] The synthesis route for PFPE(7000 g / mol)-PEG(1500 g / mol)-PFPE(7000 g / mol) was explored, utilizing copper click chemistry. This approach commenced with the production of PEG diazide and propargyl PFPE precursors. Initially, PEG was deprotonated using sodium hydroxide, facilitating the substitution of chloride in p- toluenesulfonyl chloride (pTsCI) to yield PEG ditosylate. The synthesis continued with the conversion of the tosyl group to azide using sodium azide in dimethylformamide (DMF) at 50°C.

[0472] The PFPE precursor was prepared by activating PFPE containing one terminal carboxylic acid group with oxalyl chloride, followed by immediate conversion to propargyl PFPE using propargyl amine.

[0473] The final step involved a copper-catalyzed click reaction between the prepared precursors, enhanced by the addition of neocuproine to increase reaction efficiency. This synthesis was conducted under reflux for 48 hours.

[0474] GAR-P04610W010 Application (final). docx ABK

[0475] 65

[0476] The following diblock- and driblock co-polymer surfactants have been prepared:

[0477] Table 1 : List of diblock- and driblock co-polymer surfactants

[0478] Name Di- / Triblock PEG PFPE surfactant 1 diblock 350 2000 surfactant 2 diblock 350 4000 surfactant 3 diblock 350 7000 surfactant 4 diblock 550 2000 surfactant 5 diblock 550 4000 surfactant 6 diblock 550 7000 surfactant 7 diblock 750 2000 surfactant 8 diblock 750 4000 surfactant 9 diblock 750 7000 surfactant 10 triblock 300 2000 surfactant 11 triblock 300 4000 surfactant 12 triblock 300 7000 surfactant 13 triblock 600 2000 surfactant 14 triblock 600 4000 surfactant 15 triblock 600 7000 surfactant 16 triblock 1500 2000 surfactant 17 triblock 1500 4000 surfactant 18 triblock 1500 7000

[0479] Example 2: Determination of interfacial tension

[0480] To determine the interfacial tension between the selected formulation and water, the inventors employed the pendant drop method. Initially, the formulation was diluted to a total concentration of 2.5 mM using Novec 7500 Engineered Fluid and loaded into a syringe equipped with a flat-tipped needle. The surrounding phase comprised deionized water, which was carefully introduced into a cuvette to serve as the continuous phase.

[0481] For the analysis, a drop shape analyzer was utilized. The syringe containing the surfactant formulation was securely mounted onto the analyzer, and the needle was degassed to eliminate any air bubbles that might interfere with the accuracy of the measurements. The syringe was then positioned such that the needle was vertically immersed in the water within the cuvette.

[0482] A droplet of the formulation was carefully formed at the tip of the needle, with the volume of the drop maximized to enhance the precision of the interfacial tension measurement. The observation camera was adjusted to ensure that the area of the drop

[0483] GAR-P04610W010 Application (final). docx recorded was as large as possible, thereby optimizing the accuracy of the shape analysis.

[0484] The interfacial tension was instantaneously measured by the instrument's software, which applied the Young-Laplace equation to the observed droplet shape. This equation relates the droplet shape to the interfacial tension, allowing for precise calculations based on the curvature and density differences between the two phases.

[0485] Post-formation, the drop was continuously observed for up to one hour to monitor any changes in interfacial tension over time. Data points were recorded every minute to capture any dynamic interfacial phenomena. This meticulous observation period was critical for understanding the stability and behavior of the interfacial tension in the presence of the selected formulation.

[0486] This detailed and systematic approach to measuring interfacial tension ensures high accuracy and repeatability, providing reliable data for the optimization of formulation properties in various applications. The insights gained from these measurements are integral to the formulation development process, aiding in the refinement of product performance characteristics.

[0487] Table 2 Measured IFT ranges for various surfactant formulations of the present invention comprising surfactant 1 to surfactant 18 in HFE. percentages are given with respect to the molar ratios of the surfactants. IFT ranges: A: 0-4.99 mN / m; B: 5-9.99 mN / m; C: 10-14.99 mN / m, D: 15-19.99 mN / m; E: 20-24.99 mN / m; F: 25-29.99 mN / m; G: 30-40 mN / m.

[0488] # composition IFT range

[0489] HFE G

[0490] 1 100% of surfactant 1 D

[0491] 45 50% of surfactant 1, 25% of surfactant 4, 25% of surfactant 18

[0492] D

[0493] 121 50% of surfactant 1 , 25% of surfactant 10, 25% of surfactant 11

[0494] D

[0495] 127 50% of surfactant 1, 25% of surfactant 10, 25% of surfactant 17

[0496] 25% of surfactant 1 , 25% of surfactant 2, 25% of surfactant 4, B 194 25% of surfactant 15

[0497] 314 50% of surfactant 3, 25% of surfactant 1, 25% of surfactant 15 ®

[0498] 25% of surfactant 1 , 25% of surfactant 3, 25% of surfactant 7, B

[0499] 380 25% of surfactant 11

[0500] 25% of surfactant 1, 25% of surfactant 4, 25% of surfactant 11 , A 644 25% of surfactant 13

[0501] 25% of surfactant 1, 25% of surfactant 4, 25% of surfactant 11 , B 647 25% of surfactant 16

[0502] 25% of surfactant 1, 25% of surfactant 6, 25% of surfactant 11 , B

[0503] 720 25% of surfactant 12

[0504] 755 75% of surfactant 7, 25% of surfactant 1 ®

[0505] 758 50% of surfactant 7, 25% of surfactant 1 , 25% of surfactant 10 ®

[0506] 25% of surfactant 1, 25% of surfactant 7, 25% of surfactant 11 , B 798 25% of surfactant 12

[0507] GAR-P04610W010 Application (final). docx ABK

[0508] 67

[0509] 25% of surfactant 1, 25% of surfactant 7, 25% of surfactant 13, B

[0510] 813 25% of surfactant 14

[0511] 843 50% of surfactant 8, 25% of surfactant 1, 25% of surfactant 18 B

[0512] 25% of surfactant 1, 25% of surfactant 8, 25% of surfactant 15, B

[0513] 892 25% of surfactant 18

[0514] 25% of surfactant 1, 25% of surfactant 9, 25% of surfactant 13, B

[0515] 936 25% of surfactant 16

[0516] 25% of surfactant 1 , 25% of surfactant 11 , 25% of surfactant 13, C

[0517] 1016 25% of surfactant 15

[0518] 25% of surfactant 1 , 25% of surfactant 13, 25% of surfactant 15, C

[0519] 1075 25% of surfactant 16

[0520] 1083 50% of surfactant 18, 25% of surfactant 1, 25% of surfactant 13 C

[0521] 1141 50% of surfactant 2, 25% of surfactant 3, 25% of surfactant 8 B

[0522] 1197 50% of surfactant 2, 25% of surfactant 5, 25% of surfactant 18 B

[0523] 1258 50% of surfactant 2, 25% of surfactant 11, 25% of surfactant 18 B

[0524] 1276 50% of surfactant 3, 25% of surfactant 2, 25% of surfactant 7 C

[0525] 25% of surfactant 2, 25% of surfactant 3, 25% of surfactant 4, B

[0526] 1294 25% of surfactant 10 25% of surfactant 2, 25% of surfactant 3, 25% of surfactant 4, B

[0527] 1297 25% of surfactant 12 25% of surfactant 2, 25% of surfactant 4, 25% of surfactant 5, A

[0528] 1550 25% of surfactant 12 25% of surfactant 2, 25% of surfactant 4, 25% of surfactant 8, B

[0529] 1586 25% of surfactant 12

[0530] 1593 50% of surfactant 9, 25% of surfactant 2, 25% of surfactant 4 B 25% of surfactant 2, 25% of surfactant 6, 25% of surfactant 15, B

[0531] 1717 25% of surfactant 16

[0532] 2110 50% of surfactant 3, 25% of surfactant 4, 25% of surfactant 8 B

[0533] 2119 50% of surfactant 3, 25% of surfactant 4, 25% of surfactant 16 C

[0534] 2172 50% of surfactant 3, 25% of surfactant 7, 25% of surfactant 15 B

[0535] 2198 50% of surfactant 3, 25% of surfactant 10, 25% of surfactant 11 C

[0536] 2218 50% of surfactant 3, 25% of surfactant 15, 25% of surfactant 16 C 25% of surfactant 3, 25% of surfactant 4, 25% of surfactant 5, B

[0537] 2373 25% of surfactant 16

[0538] 25% of surfactant 3, 25% of surfactant 4, 25% of surfactant 10, B

[0539] 2425 25% of surfactant 13

[0540] 2631 50% of surfactant 8, 25% of surfactant 3, 25% of surfactant 15 C

[0541] 25% of surfactant 3, 25% of surfactant 8, 25% of surfactant 14, B

[0542] 2677 25% of surfactant 16

[0543] 25% of surfactant 3, 25% of surfactant 10, 25% of surfactant 12, C

[0544] 2766 25% of surfactant 16

[0545] 2876 50% of surfactant 14, 25% of surfactant 3, 25% of surfactant 15 B

[0546] 3606 50% of surfactant 4, 25% of surfactant 5, 25% of surfactant 6 A

[0547] 3687 50% of surfactant 5, 25% of surfactant 12, 25% of surfactant 16 B

[0548] 3694 50% of surfactant 5, 25% of surfactant 13, 25% of surfactant 17 B

[0549] 3696 50% of surfactant 5, 50% of surfactant 14 B

[0550] 25% of surfactant 5, 25% of surfactant 6, 25% of surfactant 11 , B

[0551] 3770 25% of surfactant 15

[0552] 3854 25% of surfactant 5, 25% of surfactant 7, 25% of surfactant 12, B

[0553] GAR-P04610W010 Application (final). docx ABK

[0554] 68

[0555] 25% of surfactant 14

[0556] 4081 50% of surfactant 18, 25% of surfactant 5, 25% of surfactant 11 B

[0557] 4109 50% of surfactant 18, 25% of surfactant 5, 25% of surfactant 12 B

[0558] 4128 50% of surfactant 17, 25% of surfactant 5, 25% of surfactant 13 B

[0559] 4166 100% of surfactant 6 B

[0560] 4240 50% of surfactant 6, 25% of surfactant 13, 25% of surfactant 17 B

[0561] 4247 50% of surfactant 6, 50% of surfactant 15 B

[0562] 4502 50% of surfactant 11 , 25% of surfactant 6, 25% of surfactant 12 B

[0563] 4621 100% of surfactant 7 B

[0564] 4661 50% of surfactant 7, 25% of surfactant 10, 25% of surfactant 17 B

[0565] 4693 50% of surfactant 7, 50% of surfactant 16 C

[0566] 25% of surfactant 7, 25% of surfactant 8, 25% of surfactant 13, B

[0567] 4746 25% of surfactant 15

[0568] 4799 50% of surfactant 13, 25% of surfactant 7, 25% of surfactant 9 B

[0569] 25% of surfactant 7, 25% of surfactant 11, 25% of surfactant 12, C

[0570] 4877 25% of surfactant 16

[0571] 25% of surfactant 7, 25% of surfactant 12, 25% of surfactant 13, C

[0572] 4912 25% of surfactant 17

[0573] 25% of surfactant 7, 25% of surfactant 13, 25% of surfactant 14, B

[0574] 4938 25% of surfactant 17

[0575] 4985 100% of surfactant 8 B

[0576] 5048 50% of surfactant 8, 50% of surfactant 17 B

[0577] 25% of surfactant 8, 25% of surfactant 9, 25% of surfactant 10, B

[0578] 5065 25% of surfactant 14

[0579] 5103 50% of surfactant 17, 25% of surfactant 8, 25% of surfactant 9 B

[0580] 25% of surfactant 8, 25% of surfactant 10, 25% of surfactant 11 , B

[0581] 5116 25% of surfactant 12

[0582] 25% of surfactant 8, 25% of surfactant 11, 25% of surfactant 14, B

[0583] 5173 25% of surfactant 15

[0584] 25% of surfactant 8, 25% of surfactant 12, 25% of surfactant 14, B

[0585] 5201 25% of surfactant 15

[0586] 5263 50% of surfactant 16, 25% of surfactant 8, 25% of surfactant 18 C

[0587] 5268 50% of surfactant 17, 25% of surfactant 8, 25% of surfactant 18 C

[0588] 5271 100% of surfactant 9 C

[0589] 5323 50% of surfactant 9, 50% of surfactant 17 D

[0590] 5325 50% of surfactant 9, 50% of surfactant 18 C

[0591] 5376 50% of surfactant 11 , 25% of surfactant 9, 25% of surfactant 16 D

[0592] 25% of surfactant 9, 25% of surfactant 11, 25% of surfactant 12, D

[0593] 5383 25% of surfactant 16

[0594] 5455 50% of surfactant 18, 25% of surfactant 9, 25% of surfactant 13 C

[0595] 5491 100% of surfactant 10 C

[0596] 25% of surfactant 10, 25% of surfactant 13, 25% of surfactant 14, C

[0597] 5607 25% of surfactant 15

[0598] 5620 50% of surfactant 18, 25% of surfactant 10, 25% of surfactant 13 D

[0599] 5624 50% of surfactant 14, 25% of surfactant 10, 25% of surfactant 17 C 25% of surfactant 10, 25% of surfactant 14, 25% of surfactant 17, D

[0600] 5634 25% of surfactant 18

[0601] GAR-P04610W010 Application (final). docx ABK

[0602] 69

[0603] 5656 100% of surfactant 11D

[0604] 5776 100% of surfactant 12D

[0605] 5787 50% of surfactant 12, 25% of surfactant 13, 25% of surfactant 17

[0606] 5830 50% of surfactant 15, 25% of surfactant 12, 25% of surfactant 14

[0607] 5860 100% of surfactant 13 ®

[0608] 5870 50% of surfactant 13, 25% of surfactant 14, 25% of surfactant 18

[0609] 5871 50% of surfactant 13, 50% of surfactant 15

[0610] 5916 100% of surfactant 14C

[0611] 5934 50% of surfactant 15, 25% of surfactant 14, 25% of surfactant 18

[0612] 5941 75% of surfactant 16, 25% of surfactant 14

[0613] 5946 50% of surfactant 18, 25% of surfactant 14, 25% of surfactant 16

[0614] 5951 100% of surfactant 15C

[0615] 5971 100% of surfactant 16D

[0616] 5981 100% of surfactant 17

[0617] 5985 100% of surfactant 18

[0618] Example 3: General w / o droplet production

[0619] To create w / o droplets, the inventors utilized a microfluidics chip based on the design principles outlined in The et al. (The, S.-Y.; Lin, R.; Hung, L.-H.; Lee, A. P. Droplet microfluidics. Lab on a Chip 2008, 8 (2), 198-220, 10.1039 / B715524G. DOI: 10.1039 / B715524G). The microfluidics chip featured a straightforward configuration with two inlets and one outlet, enabling efficient droplet formation.

[0620] The first inlet was dedicated to the surfactant formulation, which was prepared at a total concentration of 2.5 mM in Novec 7500 Engineered Fluid. The second inlet accommodated the aqueous phase, the composition of which varied depending on the specific experimental requirements. The microchannels within the chip were precisely engineered, with a width and height of approximately 10 pm.

[0621] The design included a bifurcation of the channel carrying the surfactant formulation into two smaller channels. These channels, along with the single aqueous phase channel, converged at a T-junction. At this junction, the two surfactant formulation channels were aligned horizontally opposite each other, while the aqueous phase channel intersected them vertically. Notably, the width of all three channels tapered gradually to 5 pm as they approached the T-junction, ensuring precise control over droplet formation. The outlet channel, which also had a width of 10 pm, was positioned directly opposite the aqueous phase inlet.

[0622] To facilitate droplet production, the inlets of the microfluidic chip were connected to containers filled with the respective solutions. The solutions were introduced into the microfluidics chip using either a syringe pump or a pressure controller. The production rate of the droplets was governed by the flow rate or pressure applied, while the size of

[0623] GAR-P04610W010 Application (final). docx the droplets was determined by the ratio of the flow rates or pressures between the two phases.

[0624] The outlet channel of the microfluidic chip was connected to a collection container, where the formed w / o droplets were accumulated. This setup allowed for precise and reproducible control over droplet size and production rate, critical for various applications in formulation development and testing.

[0625] By utilizing this microfluidics approach, the inventors achieved consistent and scalable production of w / o droplets, which are essential for investigating the properties and performance of the formulations according to the present invention under controlled conditions. This methodology provides a robust platform for the systematic study and optimization of droplet-based systems in various pharmaceutical and biotechnological applications.

[0626] Example 4: Determining the viability of encapsulated cells

[0627] Unless otherwise stated, all chemicals and cell culture supplies are acquired from Sigma-Aldrich and cells are cultivated and maintained using standard and sterile cell culture conditions (37 °C and 5% CO2). To evaluate droplet stability and cell viability for cells encapsulated in droplets with surfactant formulations, we use mammalian cell lines (for example but not limited to HEK293 or CHO) cultured in DMEM cell culture media supplemented with 10% FBS. The mammalian cells are cultivated in appropriate cell culture medium in a CO2 incubator at 37°C. Upon reaching 70-80% confluency, cells are detached using trypsin-EDTA solution. The trypsin is neutralized with fresh medium, and cells are transferred to a centrifuge tube. After centrifugation at 300 x g for 5 minutes, the supernatant is discarded. The cell pellet is resuspended, and cells are counted followed by adjusting the cell concentration 1 x 10A5 cells / mL in cell culture media with 10% FBS. This cell suspension is used as the fluid for injection into the water-inlet of the droplet microfluidics device. In addition, the cell culture media can be supplemented with commercially available cell viability assays (such as Annexin V, Caspase activity assays, or Live / Dead stain, according to the manufacturer’s description) to probe cell viability using fluorescence detection approaches, as a proxy for surfactant formulation biocompatibility.

[0628] The droplet-based microfluidic device is assembled as per the description before. The device is connected to syringes filled with the cell suspension and oil phase containing the surfactant formulation according to the present invention to be investigated using appropriate tubing. The microfluidic channels are primed with the appropriate fluid to remove air bubbles. The pressurized pumps are set to desired flow rates to generate droplets (e.g., 1-10 pL / min for the aqueous phase and 10-100 pL / min for the oil phase). Droplet formation is monitored under an inverted light microscope to evaluate uniformity and stability, with small adjustments to the flow rates being made when necessary. If

[0629] GAR-P04610W010 Application (final). docx the cells are encapsulated with viability assay components doped into the cell culture media, it is also possible at this point to collect a time point for biocompatibility by adding a second parameter to the microscopy for detecting fluorescence of the cells within the droplets under flow conditions. For example, droplet parameters such as size and stability are evaluated using image analysis of bright-field illumination images followed by evaluating fluorescence signals using 488 nm and 555 nm wavelengths of light that report on the “Live / Dead” status, respectively, of the encapsulated cell. The generated droplets are collected in an Eppendorf tube for further processing.

[0630] To evaluate longer time-scales, collected droplets are transferred to a culture chamber that is gas-permeable (e.g. Ibidi cell culture chambers) and is incubated in a CO2 incubator at 37°C for the desired duration (24-72 hours). At appropriate time-points, the droplets can be reintroduced to a microfluidics flow chamber for evaluation of droplet stability and biocompatibility as described above. Alternatively, an end-point assay can be implemented where the encapsulated contents of the droplets is released using perfluoro-octanol and recovered. The released cells are then incubated with a desired viability staining solution at 37°C for 15-30 minutes and washed gently with PBS to remove excess stain. Stained cells are observed and imaged under an inverted fluorescence microscope or by flow cytometry. Images are analyzed to quantify live (green fluorescence) and dead (red fluorescence) cells.

[0631] As mentioned, the droplet stability of produced droplets using surfactant formulations is evaluated using image analysis by measuring the diameter of droplet populations over time with 0%, 1 %, and 10% FBS in the water-phase. The biocompatibility of the same surfactant formulations is analyzed compared to controls (e.g., droplets with conventional surfactants) with image analysis by measuring the fluorescence profile of cells. Statistical analyses (e.g., t-test or ANOVA) are performed to determine the significance of differences in droplet stability and cell viability.

[0632] Example 5: Determining release efficiency for the construction of lipid vesicles

[0633] To measure the release efficiency of the respective surfactant formulation, droplets with a diameter of 10 ± 1 pm were produced following the general w / o droplet production method. The aqueous phase comprised 100 pL of a solution containing 2 mM small unilamellar vesicles (SLIVs) with a composition of 30% DOBAQ, 10% DOPE, 20% DOPG, 39% DOPC, and 0.5% Liss Rhod PE, and an average diameter of approximately 120 nm. This solution also included 140 mM potassium chloride and 30 mM Tris buffer at pH 7.4. The surfactant formulation was supplemented with 10 mM of perfluoropolyether (PFPE) containing a terminal carboxylic acid group with a molecular weight of 7000 g / mol.

[0634] GAR-P04610W010 Application (final). docx Following the production of droplets, the droplet-stabilized giant unilamellar vesicles (dsGUVs), were incubated for 12 hours at 4°C to ensure stability. Excess oil was then removed, and an equal volume of release buffer (140 mM potassium chloride and 30 mM Tris buffer at pH 7.4) was added to match the inner phase. Additionally, an equal volume of a destabilizing surfactant, 1 H,1 H,2H,2H-perfluoro-1 -octanol, was introduced. The vial was sealed and gently rotated transversely about the horizontal axis to facilitate gentle mixing and increased contact area. After phase separation, the vial was stored for at least 30 minutes at 4°C.

[0635] Subsequently, 150 pL of the aqueous solution was carefully pipetted into a new Eppendorf vial, ensuring that the oil phase was not transferred. To further ensure the exclusion of oil droplets, the vial was briefly centrifuged using a table centrifuge before removing the aqueous phase. The vial was then filled up to 1000 pL with release buffer and centrifuged for one hour at 21 ,000 xg until a dye-dependent colored GUV pellet was visible. The supernatant was replaced with fresh release buffer, the pellet was resuspended, and the centrifugation process was repeated under the same conditions. Finally, the pellet was re-suspended in a total volume of 150 pL release buffer.

[0636] The release efficiency was quantified by determining the phosphate concentration in the purified GUV suspension. A calibration curve was generated using sodium dihydrogen phosphate solutions with phosphate concentrations ranging from 0.0 to 50 pg / mL in increments of 2.5 pg / mL, each in a total volume of 100 pL. To 100 pL of the GUV suspension, 200 pL of perchloric acid (70 w / v%) was added. Both the calibration samples and the GUV samples were heated to 220°C to evaporate the water. After cooling, each sample was mixed with 700 pL of an aqueous solution containing 0.45 w / v% ammonium molybdate (NH4MOO4) and 12.6 w / v% perchloric acid (HCIO4), along with 700 pL of an aqueous solution of 1.7 w / v% ascorbic acid. The mixtures were incubated for 10 minutes at 80°C, and the absorbance was measured at 820 nm.

[0637] Using the calibration curve, the lipid concentration in the GUV suspension was calculated and related to the initial 2 mM starting concentration, thereby determining the release efficiency. This method provides a precise and reliable measure of the surfactant formulation's efficiency in constructing lipid vesicles.

[0638] Example 6: Performing analytical molecular biology assays using a quantitative droplet-based pH detection assay

[0639] Quantitative analysis of pH is a fundamental measurement important for molecular biology as the pH environment of biomolecules directly impacts on their form, function, and applicability in detection assays. When attempting to transition molecular detection assays for pH from standard bulk conditions to droplet-based approaches, challenges

[0640] GAR-P04610W010 Application (final). docx emerge that are directly related to the droplet interface that is stabilized by surfactants, especially fluorosurfactants. Known challenges such as: (1) the interaction between surfactants and pH indicators leading to inaccurate pH readings due to changes in the dye's optical properties; (2) the background signal from surfactants masking the pH- dependent signal of the indicator dyes, leading to lower sensitivity and accuracy; (3) surfactants having their own buffering capacity, which affects the pH of the droplet environment independently of the sample being measured and leading to a discrepancy between the actual pH of the sample and the measured pH; and (4) the need for surfactants to be optimized not only for droplet stability in temperature-dependent and shear environments but also for compatibility with the pH indicators and the overall assay chemistry-are currently hindering the development of robust and accurate pH detection assays in droplet-based microfluidic systems. Therefore, the development and use of surfactant formulations is critical for the success of pH detection assays.

[0641] Here, the inventors developed a droplet-based pH detection assay with optimized surfactant formulations. The surfactant formulations that are inert towards the pH indicator dyes are used in droplet-based functional assays to demonstrate temperatureindependence in thermocycling the droplets and shear-resistance in flow cell environments by comparing droplet size distributions before and after droplet manipulations.

[0642] Unless otherwise noted, all chemicals are purchased from Sigma-Aldrich and microfluidics devices and flow cells are produced and operated as previously described. First, standard curves for the pH indicator dyes of interest are calibrated using buffered calibration standards injected into fluidic chambers and imaged for color or fluorescence intensity analysis using microscopy. Specifically, the pH-sensitive color change of the indicator dye in each droplet is measured using an inverted fluorescence microscope. For fluorescence measurements, the excitation and emission wavelengths are set according to the specific dye used (e.g., excitation at 488 nm and emission at 509 nm for fluorescein). The fluorescence intensity or absorbance of each droplet is recorded. This data is used to generate a calibration curve by plotting the measured signal (fluorescence or absorbance) against the known pH values of the buffer solutions.

[0643] Similarly, droplets encapsulating pH indicator dyes in strong buffering conditions (50 mM HEPES) and weak (5 mH HEPES) conditions are generated with the range of surfactant formulations of interest. These droplets are then imaged using the same microscopic set up and detectors as the standard solutions and evaluated for deviations from the expected colorimetric or fluorescence intensity expected for the given pH value. The surfactant formulations with the most stable and reproducible optical readout parameters are then tested to measure the pH of samples solutions with blinded pH

[0644] GAR-P04610W010 Application (final). docx values to evaluate the reliability of the described method. The data from the fluorescence or absorbance measurements are then compiled and analyzed. The pH values of the unknown samples are calculated by interpolating the calibration curve. Statistical analysis (e.g., standard deviation, coefficient of variation) is performed to ensure the reliability and reproducibility of the assay.

[0645] The standards and droplets are also exposed to thermocycling with 5 temperature ramps up to 65 °C followed with shear testing by reintroducing the droplets into high velocity flow conditions to evaluate temperature-dependent and shear-resistant droplet stability. After cycling and shear, the droplets are imaged and analyzed for expected colorimetric or fluorescent read out as well as droplet size distribution. Droplet size distribution is especially used as a proxy for identifying optimal surfactant formulations.

[0646] GAR-P04610W010 Application (final). docx

Claims

1. ABK75Claims1 . A formulation comprising a water-immiscible fluorocarbon solvent and at least one amphiphilic polymer, wherein the least one amphiphilic polymer is selected from a diblock co-polymer surfactant of formula A-B and a triblock co-polymer surfactant of formula B’-A’-B’, wherein A and A’ independently of each other represent a water-soluble polymer block having a weight average molecular weight in the range of 300 g / mol to 2000 g / mol, and B and B’ represent independently of each other a perfluorinated polyether block having a weight average molecular weight in the range of 1500 g / mol to 7500 g / mol, wherein the weight average molecular weights of A, A’, B, and B’ are determined by gel permeation chromatography, wherein the diblock co-polymer surfactant is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9, wherein surfactant 1 to surfactant 9 have the following formula: surfactant 1 : PFPE(2000 g / mol-L1-PEG(350 g / mol) surfactant 2: PFPE(4000 g / mol)-L1-PEG(350 g / mol) surfactant 3: PFPE(7000 g / mol)-L1-PEG(350 g / mol) surfactant 4: PFPE(2000 g / mol)-L1-PEG(550 g / mol) surfactant 5: PFPE(4000 g / mol)-L1-PEG(550 g / mol) surfactant 6: PFPE(7000 g / mol)-L1-PEG(550 g / mol) surfactant 7: PFPE(2000 g / mol)-L1-PEG(750 g / mol) surfactant 8: PFPE(4000 g / mol)-L1-PEG(750 g / mol) surfactant 9: PFPE(7000 g / mol)-L1-PEG(750 g / mol) wherein L1represents a triazole ring having the structure:O N / N .NH- / and wherein the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein surfactant 10 to surfactant 18 have the following formula:GAR-P04610W010 Application (final). docxABK76 surfactant 10: PFPE(2000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(2000 g / mol) surfactant 11 : PFPE(4000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(4000 g / mol) surfactant 12: PFPE(7000 g / mol)-L2-PEG(300 g / mol)-L3-PFPE(7000 g / mol) surfactant 13: PFPE(2000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(2000 g / mol) surfactant 14: PFPE(4000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(4000 g / mol) surfactant 15: PFPE(7000 g / mol)-L2-PEG(600 g / mol)-L3-PFPE(7000 g / mol) surfactant 16: PFPE(2000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(2000 g / mol) surfactant 17: PFPE(4000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(4000 g / mol) surfactant 18: PFPE(7000 g / mol)-L2-PEG(1500 g / mol)-L3-PFPE(7000 g / mol) wherein L2and L3represent a triazole ring having the structures:

2. The formulation according to claim 1 , wherein the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7000 (1 ,1 , 1 ,2, 2, 3, 3- heptafluoro-3-methoxy-propane), methylnonafluoro-n-butylethers, HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and1.1.1 .2.3.3-hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7200 (mixture of the isomers 1 -ethoxy-1 ,1 ,2, 2, 3, 3,4,4, 4-nonafluorobutane and 1-ethoxy-1 .1 .2.3.3.3-hexafluoro-2-(trifluoromethyl)propane), HFE-7300 (1 ,1 ,1 , 2, 2, 3, 4, 5,5,5- decafluoro-3-methoxy-4-(trifluoromethyl)pentane), HFE-7500 (3-ethoxy- 1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), FC-40(1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine), FC-43(1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N,N-bis(1 ,1 ,2,2,3,3,4,4,4-nonafluorobutyl)butan-1 - amine), FC-70 (1 ,1 ,2,2,3,3,4,4,5,5,5-undecafluoro-N,N-bis(undecafluoropentyl)- pentan-1 -amine), FC-72 (1 ,1 ,1 ,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane), perfluorodecalin, or mixtures of the afore-mentioned.

3. The formulation according to claim 1 or 2, wherein the water-immiscible fluorocarbon solvent is selected from the group consisting of HFE-7100 (mixture of the isomers 1 ,1 ,1 ,2,2,3,3,4,4-nonafluoro-4-methoxy-butane and 1 , 1 ,1 , 2,3,3- hexafluoro-3-methoxy-2-(trifluoromethyl)propane), HFE-7500 (3-ethoxy- 1 ,1 ,1 ,2,3,4,4,5,5,6,6,6-dodecafluoro-2-(trifluoromethyl)hexane), and FC-40 (1 ,1 ,2,2,3,3,4,4,4-nonafluoro-N,N-bis(nonafluorobutyl)butan-1 -amine).GAR-P04610W010 Application (final). docxAOK774. The formulation according to any one of claim 1 - 3, wherein the total concentration of the least one amphiphilic polymer is in a range between 0.5 mM x a + 0.5 mM and 10 mM x a + 10 mM, wherein a is the molar fraction of the diblock co-polymer surfactant in the least one amphiphilic polymer.

5. The formulation according to any one of claim 1 - 4, wherein the total concentration of the least one amphiphilic polymer is 2.5 mM x a + 2.5 mM, wherein a is the molar fraction of the diblock co-polymer surfactant in the least one amphiphilic polymer.

6. The formulation according to any one of claim 1 - 5, wherein the formulation comprises two amphiphilic polymers selected from a diblock co-polymer surfactant, and a triblock co-polymer surfactant, wherein the diblock-co-polymer is selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9 and the triblock co-polymer surfactant is selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein surfactant 10 to surfactant 18, wherein surfactant 1 to surfactant 18 have the meanings as defined in claim 1 .

7. The formulation according to any one of claim 1 - 5, wherein the formulation comprises three amphiphilic polymers selected from two different diblock copolymer surfactants and a triblock co-polymer surfactant or one diblock co-polymer surfactant and two different triblock co-polymer surfactants, or wherein the formulation comprises four amphiphilic polymers selected from three different diblock co-polymer surfactants and one triblock co-polymer surfactant, or four different diblock co-polymer surfactants, or two different diblock co-polymer surfactants and two different triblock co-polymer surfactants, or one diblock copolymer surfactant and three different triblock co-polymer surfactants, or four different triblock co-polymer surfactants, wherein the diblock-co-polymer surfactants are selected from the group consisting of surfactant 1 , surfactant 2, surfactant 3, surfactant 4, surfactant 5, surfactant 6, surfactant 7, surfactant 8, and surfactant 9 and the triblock co-polymer surfactants are selected from the group consisting of surfactant 10, surfactant 11 , surfactant 12, surfactant 13, surfactant 14, surfactant 15, surfactant 16, surfactant 17, and surfactant 18, wherein surfactant 10 to surfactant 18, wherein surfactant 1 to surfactant 18 have the meanings as defined in claim 1.GAR-P04610W010 Application (final). docx8. The formulation according to any one of claim 1 - 5, wherein formulation comprises the following diblock co-polymer surfactants and triblock co-polymer surfactants selected from surfactant 1 surfactant 1 , surfactant 10, surfactant 11 surfactant 1 , surfactant 10, surfactant 17 surfactant 1 , surfactant 11 , surfactant 13, surfactant 15 surfactant 1 , surfactant 13, surfactant 15, surfactant 16 surfactant 1 , surfactant 2, surfactant 4, surfactant 15 surfactant 1 , surfactant 3, surfactant 7, surfactant 11 surfactant 1 , surfactant 4, surfactant 11 , surfactant 13 surfactant 1 , surfactant 4, surfactant 11 , surfactant 16 surfactant 1 , surfactant 4, surfactant 18 surfactant 1 , surfactant 6, surfactant 11 , surfactant 12 surfactant 1 , surfactant 7, surfactant 11 , surfactant 12 surfactant 1 , surfactant 7, surfactant 13, surfactant 14 surfactant 1 , surfactant 8, surfactant 15, surfactant 18 surfactant 1 , surfactant 9, surfactant 13, surfactant 16 surfactant 10 surfactant 10, surfactant 13, surfactant 14, surfactant 15 surfactant 10, surfactant 14, surfactant 17, surfactant 18 surfactant 11 surfactant 11 , surfactant 6, surfactant 12 surfactant 11 , surfactant 9, surfactant 16 surfactant 12 surfactant 12, surfactant 13, surfactant 17 surfactant 13 surfactant 13, surfactant 14, surfactant 18 surfactant 13, surfactant 15 surfactant 13, surfactant 7, surfactant 9 surfactant 14 surfactant 14, surfactant 10, surfactant 17 surfactant 14, surfactant 3, surfactant 15 surfactant 15 surfactant 15, surfactant 12, surfactant 14 surfactant 15, surfactant 14, surfactant 18 surfactant 16 surfactant 16, surfactant 14 surfactant 16, surfactant 8, surfactant 18 surfactant 17 surfactant 17, surfactant 5, surfactant 13 surfactant 17, surfactant 8, surfactant 18 surfactant 17, surfactant 8, surfactant 9 surfactant 18 surfactant 18, surfactant 1 , surfactant 13 surfactant 18, surfactant 10, surfactant 13 surfactant 18, surfactant 14, surfactant 16 surfactant 18, surfactant 5, surfactant 11 surfactant 18, surfactant 5, surfactant 12 surfactant 18, surfactant 9, surfactant 13GAR-P04610W010 Application (final). docxsurfactant 2, surfactant 11 , surfactant 18 surfactant 2, surfactant 3, surfactant 4, surfactant 10 surfactant 2, surfactant 3, surfactant 4, surfactant 12 surfactant 2, surfactant 3, surfactant 8 surfactant 2, surfactant 4, surfactant 5, surfactant 12 surfactant 2, surfactant 4, surfactant 8, surfactant 12 surfactant 2, surfactant 5, surfactant 18 surfactant 2, surfactant 6, surfactant 15, surfactant 16 surfactant 3, surfactant 1 , surfactant 15 surfactant 3, surfactant 10, surfactant 11 surfactant 3, surfactant 10, surfactant 12, surfactant 16 surfactant 3, surfactant 15, surfactant 16 surfactant 3, surfactant 2, surfactant 7 surfactant 3, surfactant 4, surfactant 10, surfactant 13 surfactant 3, surfactant 4, surfactant 16 surfactant 3, surfactant 4, surfactant 5, surfactant 16 surfactant 3, surfactant 4, surfactant 8 surfactant 3, surfactant 7, surfactant 15 surfactant 3, surfactant 8, surfactant 14, surfactant 16 surfactant 4, surfactant 5, surfactant 6 surfactant 5, surfactant 12, surfactant 16 surfactant 5, surfactant 13, surfactant 17 surfactant 5, surfactant 14 surfactant 5, surfactant 6, surfactant 11 , surfactant 15 surfactant 5, surfactant 7, surfactant 12, surfactant 14 surfactant 6 surfactant 6, surfactant 13, surfactant 17 surfactant 6, surfactant 15 surfactant 7 surfactant 7, surfactant 1 surfactant 7, surfactant 1 , surfactant 10 surfactant 7, surfactant 10, surfactant 17 surfactant 7, surfactant 11 , surfactant 12, surfactant 16 surfactant 7, surfactant 12, surfactant 13, surfactant 17 surfactant 7, surfactant 13, surfactant 14, surfactant 17 surfactant 7, surfactant 16 surfactant 7, surfactant 8, surfactant 13, surfactant 15 surfactant 8 surfactant 8, surfactant 1 , surfactant 18 surfactant 8, surfactant 10, surfactant 11 , surfactant 12 surfactant 8, surfactant 11 , surfactant 14, surfactant 15 surfactant 8, surfactant 12, surfactant 14, surfactant 15 surfactant 8, surfactant 17 surfactant 8, surfactant 3, surfactant 15 surfactant 8, surfactant 9, surfactant 10, surfactant 14 surfactant 9 surfactant 9, surfactant 11 , surfactant 12, surfactant 16 surfactant 9, surfactant 17 surfactant 9, surfactant 18 surfactant 9, surfactant 2, surfactant 4.GAR-P04610W010 Application (final). docx9. The formulation according to any one of claim 1 - 5, wherein formulation comprises the following diblock co-polymer surfactants and triblock co-polymer surfactants selected from surfactants molar ratio surfactant 1 surfactant 1 , surfactant 10, surfactant 11 2:1 :1 surfactant 1 , surfactant 10, surfactant 17 2:1 :1 surfactant 1 , surfactant 2, surfactant 4, surfactant 15 1 : 1 : 1 : 1 surfactant 1 , surfactant 3, surfactant 7, surfactant 11 1 : 1 : 1 : 1 surfactant 1 , surfactant 4, surfactant 11 , surfactant 13 1 : 1 : 1 : 1 surfactant 1 , surfactant 4, surfactant 11 , surfactant 16 1 : 1 : 1 : 1 surfactant 1 , surfactant 4, surfactant 18 2:1 :1 surfactant 1 , surfactant 6, surfactant 11 , surfactant 12 1 : 1 : 1 : 1 surfactant 1 , surfactant 7, surfactant 11 , surfactant 12 1 : 1 : 1 : 1 surfactant 1 , surfactant 7, surfactant 13, surfactant 14 1 : 1 : 1 : 1 surfactant 1 , surfactant 8, surfactant 15, surfactant 18 1 : 1 : 1 : 1 surfactant 1 , surfactant 9, surfactant 13, surfactant 16 1 : 1 : 1 : 1 surfactant 11 , surfactant 6, surfactant 12 2:1 :1 surfactant 13 surfactant 13, surfactant 7, surfactant 9 2:1 :1 surfactant 14, surfactant 3, surfactant 15 2:1 :1 surfactant 17, surfactant 5, surfactant 13 2:1 :1 surfactant 17, surfactant 8, surfactant 9 2:1 :1 surfactant 18, surfactant 5, surfactant 11 2:1 :1 surfactant 18, surfactant 5, surfactant 12 2:1 :1 surfactant 2, surfactant 11 , surfactant 18 2:1 :1 surfactant 2, surfactant 3, surfactant 4, surfactant 10 1 : 1 : 1 : 1 surfactant 2, surfactant 3, surfactant 4, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 3, surfactant 8 2:1 :1 surfactant 2, surfactant 4, surfactant 5, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 4, surfactant 8, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 5, surfactant 18 2:1 :1 surfactant 2, surfactant 6, surfactant 15, surfactant 16 1 : 1 : 1 : 1 surfactant 3, surfactant 1 , surfactant 15 2:1 :1 surfactant 3, surfactant 4, surfactant 10, surfactant 13 1 : 1 : 1 : 1 surfactant 3, surfactant 4, surfactant 5, surfactant 16 1 : 1 : 1 : 1 surfactant 3, surfactant 4, surfactant 8 2:1 :1 surfactant 3, surfactant 7, surfactant 15 2:1 :1 surfactant 3, surfactant 8, surfactant 14, surfactant 16 1 : 1 : 1 : 1 surfactant 4, surfactant 5, surfactant 6 2:1 :1 surfactant 5, surfactant 12, surfactant 16 2:1 :1 surfactant 5, surfactant 13, surfactant 17 2:1 :1 surfactant 5, surfactant 14 1 :1 surfactant 5, surfactant 6, surfactant 11 , surfactant 15 1 : 1 : 1 : 1 surfactant 5, surfactant 7, surfactant 12, surfactant 14 1 : 1 : 1 : 1 surfactant 6GAR-P04610W010 Application (final). docxABK81 surfactant 6, surfactant 13, surfactant 17 2:1 :1 surfactant 6, surfactant 15 1 :1 surfactant 7 surfactant 7, surfactant 1 3:1 surfactant 7, surfactant 1 , surfactant 10 2:1 :1 surfactant 7, surfactant 10, surfactant 17 2:1 :1 surfactant 7, surfactant 13, surfactant 14, surfactant 17 1 : 1 : 1 : 1 surfactant 7, surfactant 8, surfactant 13, surfactant 15 1 : 1 : 1 : 1 surfactant 8 surfactant 8, surfactant 1 , surfactant 18 2:1 :1 surfactant 8, surfactant 17 1 :1 surfactant 8, surfactant 10, surfactant 11 , surfactant 12 1 : 1 : 1 : 1 surfactant 8, surfactant 11 , surfactant 14, surfactant 15 1 : 1 : 1 : 1 surfactant 8, surfactant 12, surfactant 14, surfactant 15 1 : 1 : 1 : 1 surfactant 8, surfactant 9, surfactant 10, surfactant 14 1 : 1 : 1 : 1 surfactant 9, surfactant 2, surfactant 4 2:1 :

110. The formulation according to any one of claim 1 - 5, wherein formulation comprises the following diblock co-polymer surfactants and triblock co-polymer surfactants selected from surfactants molar ratio surfactant 1 surfactant 2, surfactant 4, surfactant 5, surfactant 12 1 : 1 : 1 : 1 surfactant 2, surfactant 4, surfactant 8, surfactant 12 1 : 1 : 1 : 1 surfactant 4, surfactant 5, surfactant 6 2:1 :1.

11. The formulation according to any one of claims 1 - 10, wherein the formulation is devoid of free perfluoropolyether carboxylic acid.

12. The formulation according to any one of claims 1 - 10, further comprising 0.1 mM to 15 mM of a perfluoropolyether carboxylic acid.

13. The formulation according to any one of claims 1 - 12, wherein the at least one amphiphilic polymer has a purity of at least 95% as determined by NMR spectroscopy.

14. The formulation according to any one of claims 1 - 12, wherein the at least one amphiphilic polymer has a purity of at least 98% as determined by NMR spectroscopy.GAR-P04610W010 Application (final). docx

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