WO2019110078A1 - Uv curable mounting medium - Google Patents

Uv curable mounting medium Download PDF

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Publication number
WO2019110078A1
WO2019110078A1 PCT/EP2017/001407 EP2017001407W WO2019110078A1 WO 2019110078 A1 WO2019110078 A1 WO 2019110078A1 EP 2017001407 W EP2017001407 W EP 2017001407W WO 2019110078 A1 WO2019110078 A1 WO 2019110078A1
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WO
WIPO (PCT)
Prior art keywords
mounting medium
medium according
mounting
ebecryl
curable
Prior art date
Application number
PCT/EP2017/001407
Other languages
French (fr)
Inventor
Matthias STOLMÁR
Alexander Schmid
Original Assignee
Orsatec Gmbh
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Orsatec Gmbh filed Critical Orsatec Gmbh
Priority to PCT/EP2017/001407 priority Critical patent/WO2019110078A1/en
Publication of WO2019110078A1 publication Critical patent/WO2019110078A1/en

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08F—MACROMOLECULAR COMPOUNDS OBTAINED BY REACTIONS ONLY INVOLVING CARBON-TO-CARBON UNSATURATED BONDS
    • C08F290/00—Macromolecular compounds obtained by polymerising monomers on to polymers modified by introduction of aliphatic unsaturated end or side groups
    • C08F290/08—Macromolecular compounds obtained by polymerising monomers on to polymers modified by introduction of aliphatic unsaturated end or side groups on to polymers modified by introduction of unsaturated side groups
    • C08F290/14—Polymers provided for in subclass C08G
    • C08F290/147—Polyurethanes; Polyureas
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
    • C08G18/00—Polymeric products of isocyanates or isothiocyanates
    • C08G18/06—Polymeric products of isocyanates or isothiocyanates with compounds having active hydrogen
    • C08G18/08—Processes
    • C08G18/16—Catalysts
    • C08G18/22—Catalysts containing metal compounds
    • C08G18/24—Catalysts containing metal compounds of tin
    • C08G18/244—Catalysts containing metal compounds of tin tin salts of carboxylic acids
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
    • C08G18/00—Polymeric products of isocyanates or isothiocyanates
    • C08G18/06—Polymeric products of isocyanates or isothiocyanates with compounds having active hydrogen
    • C08G18/28—Polymeric products of isocyanates or isothiocyanates with compounds having active hydrogen characterised by the compounds used containing active hydrogen
    • C08G18/67—Unsaturated compounds having active hydrogen
    • C08G18/6795—Unsaturated polyethers
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
    • C08G18/00—Polymeric products of isocyanates or isothiocyanates
    • C08G18/06—Polymeric products of isocyanates or isothiocyanates with compounds having active hydrogen
    • C08G18/70—Polymeric products of isocyanates or isothiocyanates with compounds having active hydrogen characterised by the isocyanates or isothiocyanates used
    • C08G18/72—Polyisocyanates or polyisothiocyanates
    • C08G18/74—Polyisocyanates or polyisothiocyanates cyclic
    • C08G18/76—Polyisocyanates or polyisothiocyanates cyclic aromatic
    • C08G18/7657—Polyisocyanates or polyisothiocyanates cyclic aromatic containing two or more aromatic rings
    • C08G18/7664—Polyisocyanates or polyisothiocyanates cyclic aromatic containing two or more aromatic rings containing alkylene polyphenyl groups
    • C08G18/7671—Polyisocyanates or polyisothiocyanates cyclic aromatic containing two or more aromatic rings containing alkylene polyphenyl groups containing only one alkylene bisphenyl group
    • C—CHEMISTRY; METALLURGY
    • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
    • C09D—COATING COMPOSITIONS, e.g. PAINTS, VARNISHES OR LACQUERS; FILLING PASTES; CHEMICAL PAINT OR INK REMOVERS; INKS; CORRECTING FLUIDS; WOODSTAINS; PASTES OR SOLIDS FOR COLOURING OR PRINTING; USE OF MATERIALS THEREFOR
    • C09D4/00—Coating compositions, e.g. paints, varnishes or lacquers, based on organic non-macromolecular compounds having at least one polymerisable carbon-to-carbon unsaturated bond ; Coating compositions, based on monomers of macromolecular compounds of groups C09D183/00 - C09D183/16
    • C09D4/06—Organic non-macromolecular compounds having at least one polymerisable carbon-to-carbon unsaturated bond in combination with a macromolecular compound other than an unsaturated polymer of groups C09D159/00 - C09D187/00

Definitions

  • Mounting media are used in biology and medicine, especially in the areas of histology, histochemistry, immunology and cytochemistry to fix a biological samples for microscopy to make them suitable for long storage.
  • the samples for examples tissue or blood samples or cells are placed on a microscope slide, typically made of glass and finally covered with a cover glass.
  • the cover glasses usually fixed with a mounting medium on the microscopic slide. In one aspect this improves the usability.
  • the fixed sample can be stored over many years without degradation, since the sample is emerged in the mounting media and is therefore protected against humidity, oxygen and light.
  • a mounting media is a fast curing and a high inertness after curing in reactions with water or alcohol. Further, it should be transparent and should have a refractive index which is comparable to the cover glass.
  • the gold standard for the fixation of microscopical samples is“EUKITT ® ” (ORSAtec), Germany), a xylene based mounting medium. “EUKITT ® ” cures fast, is completely transparent and has a very small loss on curing.
  • a certain disadvantage is the use of xylene which is in small amount and by inhalation toxic and requires upon handling the use of a protective mask.
  • Further mounting media in the prior art use limonene instead of xylene. A limonene based mounting medium yield also well conserved tissues. However, limonene can lead to skin irritations and has further a very strong smell which may be regarded as irritating upon daily use.
  • a third group of mounting media is based on UV-curable polyacrylates. These offer the advantage that the mounted sample can be covered after a comparatively short exposure to UV irradiation with a coating of polyacrylate.
  • UV-curable mounting media An example for a UV-curable mounting media is CureMount II (Electronic Microscopy Sciences, USA), which is based on isobornyl-acrylate as acrylate-monomer. However, this is a toxic monomer which exposes its users to health risks due to its volatility.
  • CureMount II Electronic Microscopy Sciences, USA
  • This is a toxic monomer which exposes its users to health risks due to its volatility.
  • the presentation “Energy Curable Oligomers for Outdoor Applications” (www.mycoatingresins.com Q2 2013) gives an overview on UV curable Oligomers.
  • US 3,467,617 discloses a UV curable mounting media which is non-fluorescent.
  • EP 0104825 discloses in a general manner a curable adhesive composition, which has after curing a refractive index substantially similar to that of the sample.
  • a preferred composition comprises a diacrylate-terminated polyurethane, a diacrylate ester of an epoxy resin and a diethoxyacetophenone initiator. It is formed on the mounting surface as a solution in a conventional organic solvent, such as isopropanol or toluene.
  • the UV curable mounting media of the prior art are usually a three component systems in which is difficult to regulate the viscosity. Is the mounting media to viscous it might form bubbles upon its application to the microscope slide which will make the subsequent microscopic analysis more difficult or even impossible. Further, the UV curable mounting media usually are based on monomers which are partial toxic and highly volatile.
  • VOC Volatile Organic Compounds
  • a further requirement is the selective removal of the cover glass by an aprotic non-polar solvent to carry out a necessary staining or subsequent tests on the mounted sample.
  • a UV curable mounting medium comprising a composition containing mono and/or bifunctional short chain acrylate oligomers or a mixture of mono- and a bifunctional short chain acrylate oligomer(s) with a molecular weight in the range from 100 to 1000 and a photopolymerisation initiator, wherein the photopolymerisation initiator is present in an amount of 0.81 to 3.5 wt%.
  • a cover glass which is mounted with the mounting medium according to the invention can be removed selectively after curing with xylene or toluene.
  • the cover glass can be removed by an aprotic, non-polar solvent. Therefore, the composition according to the invention provides the easy possibility to allow additional staining or subsequent tests on the mounted tissue. Strong polar solvents, like acetone or acetic acid do not only remove a mounted sample from the cover glass but also from the mounting slide.
  • the oligomer of the composition according to the present invention contains higher functional units, then a three-dimensional network is formed which cannot be removed by xylene, i.e. a removal of only the cover glass is impossible.
  • the amount of photopolymerisation initiator is below 0.89 wt%, the mounting medium does not harden within the standard hardening time of 30 to 120 seconds upon exposure to a UV/VIS light source, for example a commercial light source UV or LED (1 to 100 watt) with a maximum wavelength of 365 nm). Is the initiator present in an amount of more than 3.5 wt%, the cover glass cannot be remove with xylene at all even within 14 days.
  • the cover glass can be removed in a time span of more than 24 hours.
  • the most preferred amount of photopolymerisation initiator is therefore in the range of 0.89, preferably 0.90 to 1.95 wt% which allows the selective removal of the cover glass after curing by a non-polar aprotic solvent within a commercially reasonable time, i.e. within 1 to 2 hours.
  • a further advantage of the mounting medium according to the present invention is the compatibility with washing solvents like alcohols as for example ethanol, isopropanol, and propanol, isoparaffines and petrol ether.
  • the tissue is usually washed with a sequence of several alcohols and before mounting washed with nonpolar non protic solvents to provide a compatibility with a mounting media and to remove the last rest of water.
  • the present mounting media can also be applied directly on the samples or tissue cuts which are immersed in pure isopropanol or ethanol. This shortens the entire washing process and consumes less chemical solvents.
  • the mounting medium according to the present invention shows a very good compatibility with xylene alternatives, like isoparaffines or petrol ethers or benzene fractions containing compounds of small chain lengths (for example obtainable under the trademark Tissue Clear ® from Sakura).
  • xylene alternatives like isoparaffines or petrol ethers or benzene fractions containing compounds of small chain lengths (for example obtainable under the trademark Tissue Clear ® from Sakura).
  • cover slipper machines an advantage of the use of the mounting medium according to the present invention is that the sample cannot dry in the cover slipper machine even upon staying a longer life span in the machine or during a malfunction when the tissue is washed before mounting with the mono and/or bifunctional acrylic oligomer or mixtures thereof.
  • a still further advantage of the mounting medium according to the invention is that a significantly lower amount of mounting medium can be used compared to physically curable mounting media in the prior art: only 50% of the amount of a mountain medium of the prior is necessary when using a mounting medium according to the invention to obtain the same effects and protection of the samples
  • the mounting medium has only at least one monofunctional short chain acrylate oligomers, i.e. monofunctional reactive units.
  • the mounting media contains only at least bifunctional short chain acrylate oligomers, i.e. bifunctional units.
  • the mounting media comprises a mixture of at least one mono and at least one bifunctional short chain acrylate oligomers, i.e. mono and bifunctional reactive units.
  • the mounting medium according to the invention has preferably after curing a refractive index of 1.3 to 1.7. It is important that the refractory index of the mounting media is close to the refractory index of the cover glass and/or the mounting slide so that the resolution during analysis by a microscope is very good, to detect and determine very fine tissue structures etc. Is the refractory index higher or lower than the above mentioned limits, the resolution is markedly lower even if the contrast might be higher. If the mounting media has the same refractory index as the glass and is also in the range of the sample then, the sample would no longer be visible due to extinction of the wavelengths.
  • the refractory index of a non stained histological sample is in the range of 1 .53 to 1.54 and a refractory index of a cover glass is around 1 .48 to 1.52 depending on the manufacturer Therefore, in a most preferred embodiment of the invention, a refractory index of the cured mounting medium is in the range of 1.47 to 1.55. Only for specific applications in microscopy (diatomaceous preparation, a refractory index of up to 1.7 is preferred (yellow medium). The lowest refractory index is 1.3 to obtain good results. Below this limit, no satisfactory resolution is possible.
  • a mounting medium comprises a mixture/composition of the following constituents:
  • Aliphatic urethane acrylates are all derivatives of urethanacrylates whose side chains consist of saturated or unsaturated branched or non-branched or cyclic aliphatic rests.
  • the use of multifunctional urethane acrylates is possible, i. e. of urethane acrylate based compounds with more than one acryl group. Therefore, the aliphatic urethane acrylate according to the invention can comprise 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8 or at least 9 acryl groups.
  • the aliphatic urethane acrylate comprises two or more acryl groups.
  • the aliphatic urethane acrylate has exactly two acryl groups.
  • the molecular weight of the aliphatic urethane acrylate according to the invention is in the range of 800 to 2000 g/mol, preferably between 1000 and 1500 g/mol and especially prefer between 1000 and 1200 g/mol.
  • Ebecryl 8402 Ebecryl 265, Ebecryl 837, Ebecryl 8254, Ebecryl 8409, Ebecryl 8465, Ebecryl 1291 , Ebecryl 1290 or Ebecryl 1259 (all available by Allnex, Belgium).
  • the mounting medium according to the invention comprises in an embodiment 40 wt% to 80 wt% and to 80 wt%, preferably 50 wt% to 70 wt%, most preferred 55 wt% to 65 wt% in specific embodiments 60 wt%, 63 wt% or 65 wt% of the aliphatic urethane acrylate. Is the amount of the aliphatic urethane acrylate to low, that is especially lower than 40 wt%, then, the mounting medium according to the invention is less resistant to pressure after curing and may crack.
  • the mounting media according to the invention comprises a short chain acrylate oligomer.
  • short chain acrylate oligomer in the sense of the present invention means a acrylate oligomer with a chain length of 2 to 9 acrylic acid units (monomers) wherein basically all chain length, i. e. chains comprising of 2, 3, 4, 5, 6, 7, 8 and 9 monomers are suitable for carrying out the present invention especially preferred oligomers consisting of 3 to 8 monomers.
  • the side chains of the short chain acrylate oligomers can also be modified saturated or unsaturated, branched or unbranched or cyclic aliphatic rests.
  • the short chain acrylate oligomer comprises three or more acryl groups (monomers).
  • the short chain acrylate oligomer comprises exactly three acryl groups (monomers).
  • the term also comprises molecules with only one acryl group linked to a monomeric aliphatic or olefin containing chain or a“spacer” chain between two acrylic groups, wherein the chain and the“spacer” chain are modified saturated or unsaturated, branched or unbranched or cyclic aliphatic rests.
  • the molecular weight of the short chain acrylate oligomer according to the invention is between 150 to 1000 g/mol, preferably between 300 und 800 g/mol, an most referred between 450 to 600 g/mol.
  • Ebecryl LEO 10501 Ebecryl 130, Ebecryl 140, Ebecryl 150 or Ebecryl 160
  • Ebecryl 1 13, Ebecryl 114, Ebecryl 11 or HDDA especially preferred are Ebecryl 1 13, Ebecryl 114, Ebecryl 11 and HDDA (1 ,6-hexanediol diacrylate) (all available from Allnex, Belgium).
  • the mounting medium according to the invention comprises 20 wt% to 60 wt%, preferably 30 wt% to 50 wt%, most preferred 25 wt%, 30 wt%, 35 wt% or 40 wt% of the short chain acrylate. Is the amount of aliphatic urethane acrylate too low, the viscosity of the mounting media according to the invention is too high which is difficult in handling and prevents its use in cover slipper machines.
  • the aliphatic urethane acrylate and/or the short chain acrylate oligomer are multifunctional. In a preferred embodiment, both components are multifunctional. In a further preferred embodiment, the aliphatic urethane acrylate is bifunctional or trifunctional and the short chain acrylate oligomer is biifunctional.
  • photopolymerisation initiator all compounds can be used in the present invention which form during UV irradiation radicals.
  • photopolymerisation initiators from the group of acetophenones, benzyl or benzoine compounds, benzophenones, cationic photopolymerisation initiators or thioxanthone alone or in combination with one another can be used.
  • a photopolymerisation initiator commercially available compounds can be used alone or in combination thereof, for example Omnirad 184, Omnirad 500, Omnirad 1173, Omnirad 2959, Omnirad 819, Omnirad MBF or Omnirad TPO (all available from IGM Resins, Netherlands). Especially preferred is Omnirad 1 173 alone. IGM Resins has taken over production of photopolymerisation initiators from BASF with former Tradename Irgacure.
  • the mounting medium according to the invention comprises 0.5 to 3.5 wt% of the photopolymerisation initiator.
  • the amounting medium according to the invention comprises 0.9 to 1.95 wt%, especially 1.0 to 3.5 wt%, especially 1.0 to 1.95 wt% of the photopolymerisation initiator as already discussed above.
  • the mounting medium according to the invention comprises ca. 0.1 wt%, 1.0 wt%, 1.2 wt%, 1.3 wt%, 1.4 wt%, 1.5 wt%, 1.6 wt%, 1 .7 wt%, 1.8 wt% of the photopolymerisation initiator.
  • the photopolymerisation initiator of the mounting medium according to the invention is a short or long wave photopolymerisation initiator or a mixture thereof.
  • the mounting medium according to the invention can be adapted to different areas of use and UV sources.
  • the mounting medium according to the invention can be polymerized with a multitude of various UV sources.
  • the viscosity of mounting media can be measured by different tools.
  • a flow cap-system is used (according DIN 53 21 1 ).
  • the ideal viscosity for a classic system like EUKITT® (physically drying) is around 120 sec flow time in DIN cap 4 (at 20°C).
  • EUKITT® physically drying
  • the ideal viscosity range of UV-curable mounting media according to the present invention is between 15 seconds and 90 seconds in a DIN 4 flow cap system at 20°C.
  • the mounting medium according to the invention has a preferred viscosity in the range of 20 to 70 sec flow time in DIN cap 4 (at . 20°C), still more preferred 40-60 and in further specific embodiments 45, 50 and 55 sec flow time in DIN cap 4 (at 20°C). This provides in one aspect an easier handling and in another aspect the formation of bubbles during mounting is avoided.
  • the mounting medium according to the invention is easy to store and can also be used by applying usual security measures in chemical laboratories.
  • the mounting medium according to the invention contains only a small amount of highly volatile toxic compounds and avoids therefore the application of specific security provisions. Upon irradiation with UV light, it is hardening very fast and without a shrinking. After hardening, it is completely transparent and can be stored over a long period of 1 to 15 years without discolor or forming cracks Due to its viscosity it is also advantageously useable for cover slipper machines. It is possible to obtain a mounted sample without forming bubbles and the mounting medium according to the invention conserve the samples without altering its structure or color. Further, the mounted samples are protected against humidity, oxygen, day light and UV light and further external influences and protect the samples against degradation.
  • the mounting medium according to the invention has positive polarization properties and shows no polarization colors or lines on mounting slides and cover glasses.
  • absorption maximum (UV/VIS) of a substance is its absorption maximum in the range of UV and visible light.
  • the absorption maximum (UV/VIS) of a substance determines its color.
  • Transparent substances have an absorption maximum (UV/VIS) of less than 380 nm, yellow substances an absorption maximum of 440 nm. Substances which do not alter its absorption maximum substantially even after several years do not alter its color and do not show a yellowing. A non-substantial change of absorption maximum is present if the absorption maximum changes within three years less than 30 nm.
  • the mounting medium according to the invention is characterized in that it is colorless after hardening/curing and the difference in the absorption maximum measured after one day after hardening/curing and after exactly three years after hardening/curing is less than 30 nm.
  • the mounting medium according to the invention has even after a long storage no or only a small staining effect and is therefore also extremely suitable for the mounting of samples which have to be examined over a long period of time on several occasions.
  • a mounting medium has also to have a specific refractive index after curing. This refractive index is to be adapted to the mounted tissue as also to the microscope objective with which the sample is examined.
  • the refractory index of the mounting medium can be influenced by its chemical composition. Especially by mixing different chemical compounds, a desired refractory index can be obtained.
  • the person skilled in the art is well aware of methods for the measuring of the indetermination of the refractory index, for example by use of a refractometer.
  • the refractory index of the mounting medium is after curing in the range from 1.3 to 1.7.
  • the mounting medium of the present invention can be manufactured by processes known essentially to a person skilled in the art.
  • the mounting medium according to the invention can be obtained by mixing of the separate components. Either all of the components can be mixed together or two components are mixed in a first step and all the other components can be added in one or more steps.
  • composition for the mounting medium can be used as a nail polish or as a glue.
  • the mounting medium of the present invention can comprise further components for example inert solvents, diluents, indicators, light stabilizers, antioxidants, pigments or colors.
  • the mounting medium according to the invention can be used as any other mounting medium known in the art.
  • Biological samples which have to be mounted with the mounting medium according to the invention comprise cells, bacteria, tissue cuts, tissue samples, blood, plasma or serum samples.
  • the samples can either be directly mounted or previously stained.
  • the staining methods are known essentially known to a person skilled in the art and comprise for example hematoxyline-eosin staining, Giemsa staining, gram staining and toluidine staining.
  • the mounting medium according to the invention can be used for the mounting of flat samples liker cells or tissue cuts or mounting of three dimensional structures as tissues, insects, plants or pieces thereof. Further, the mounting medium according to the invention is especially suitable for the mounting of samples with low contrast or colorless samples.
  • the mounting medium according to the invention can be hardened/cured by any conventional UV source, for example with UV lamps or UV hardening devices.
  • the irradiation time depends on the exact composition of the mounting medium according to the invention. A person skilled in the art is aware of how to determine the optimum irradiation time and intensity.
  • UV lights source with a wave length range of 220 to 360 nm during 120 sec. are used for curing at the mounting medium according to the invention.
  • the mounting medium according to the invention hardened after applying UV irradiation within 120 sec., preferably within 30 sec.
  • the mounting medium according to the invention is also suitable for the mounting of fluorescing and fluorescing stained samples.
  • the mounting medium according to the invention contains an anti-bleaching agent like BHT (Butylhydroxytoluol), Irganox 1076 (Octadecyl 3-(3,5-di-tert-butyl-4-hydroxyphenyl) propionate), Irganox 1010 (Pentaerythrittetrakis (3-(3,5-di-tert-butyl-4-hydroxyphenyl) propionate), Tinuvin 312 (N-(2-Ethoxyphenyl)-N’-(4-thylphenyl)-ethlyene diamid), Tinuvin 1577 (2-(4,6-diphenyl-1 ,3,5-triazin-2-yl)-5-(hexyl)oxyphenol) and other suitable agents alone or in combination thereof which avoid a bleaching of the fluorescence staining agent in the sample.
  • BHT
  • Irgacure 1 173 (2.862 g (1.565 wt%) is mixed with a dissolver
  • Irgacure 1173 (1 .653 g (0.90 wt%) was mixed with a dissolver
  • the mixture was then mixed during 15 minutes at room temperature and bottled and
  • Irgacure 1 173 (2.40 g (1.216 wt%) was mixed with a dissolver
  • the mixture was then mixed during 15 minutes at room temperature and bottled and
  • the mixture wass then mixed during 15 minutes at room temperature and bottled and stored in an aluminium bottle.
  • Ebecryl 8402 were placed in a 500 ml flask and heated in a water bath at 60° C by mixing (IKA Eurostar 200 P4 Control) with a dissolver disk (diameter 6 cm).
  • IKA Eurostar 200 P4 Control IKA Eurostar 200 P4 Control
  • Ebecryl LEO 10501 were dissolved and mixed and heated in a water bath at 60° C with a dissolver disk (diameter 3 cm) and with a IKA RW 27 D mixer.
  • Ebecryl 1259 were placed in a 500 ml flask and heated in a water bath at 60° C by mixing (IKA Eurostar 200 P4 Control) and mixed with a dissolver disk (diameter 6 cm).
  • a second 250 ml flask 5.0 (1.87 wt%) Darocur TPO and 2.0 g (0.75 wt%) Darocur MBF in 80.4 g g (30.03 wt%)
  • Ebecryl LEO 10501 was dissolved under mixing and heating in water bath at 60° C with a dissolver disk (diameter 3 cm) and a IKA RW 27 D mixer
  • Ebecryl 265 were placed in a 500 ml flask and heated in a water bath at 60° C by mixing (IKA Eurostar 200 P4 Control) and mixed with a dissolver disk (diameter 6 cm).
  • IKA Eurostar 200 P4 Control In a second 250 ml flask, 8.75 (2.5 wt%) Irgacure 184 in 139.48 g (39.85 wt%) Ebecryl LEO 140 was dissolved by mixing and heating in water bath at 60° C with a dissolver plate (diameter 3 cm) and IKA RW 27 D mixer.
  • Ebecryl 160 g (44.0 wt%) Ebecryl 265 were placed in a 500 ml flask and heated in a water bath at 60° C by mixing (IKA Eurostar 200 P4 Control) and mixed with a dissolver disk (diameter 6 cm).
  • IKA Eurostar 200 P4 Control In a second 250 ml flask, 6.55 (1.8 wt%) Irgacure 819 in 197.1 g (54.2 wt%) Ebecryl 160 was dissolved under mixing and heating in water bath at 60° C with a dissolver disk (diameter 3 cm) and IKA RW 27 D mixer.
  • the Irgacure 819 which was completely dissolved in Ebecryl 160 was added to the preheated Ebecryl 265. The mixture was mixed for further 15 minutes and bottled and stored in an aluminum bottle.

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Abstract

The present invention relates to a UV curable mounting medium which can be used for the mounting of biological samples for microscopy and histology. The mounting medium according to the present invention comprises a short chain acrylate oligomer with one and/or two reactive functional groups and a photopolymerisation initiator in an amount of 0.89 to 3.5 wt%.

Description

UV-curable mounting medium
Mounting media are used in biology and medicine, especially in the areas of histology, histochemistry, immunology and cytochemistry to fix a biological samples for microscopy to make them suitable for long storage. Usually, the samples, for examples tissue or blood samples or cells are placed on a microscope slide, typically made of glass and finally covered with a cover glass. To avoid a movement of slide and cover glass, the cover glasses usually fixed with a mounting medium on the microscopic slide. In one aspect this improves the usability. Further, the fixed sample can be stored over many years without degradation, since the sample is emerged in the mounting media and is therefore protected against humidity, oxygen and light.
One of the requirements of a mounting media is a fast curing and a high inertness after curing in reactions with water or alcohol. Further, it should be transparent and should have a refractive index which is comparable to the cover glass. The gold standard for the fixation of microscopical samples is“EUKITT®” (ORSAtec), Germany), a xylene based mounting medium. “EUKITT®” cures fast, is completely transparent and has a very small loss on curing. A certain disadvantage is the use of xylene which is in small amount and by inhalation toxic and requires upon handling the use of a protective mask. Further mounting media in the prior art use limonene instead of xylene. A limonene based mounting medium yield also well conserved tissues. However, limonene can lead to skin irritations and has further a very strong smell which may be regarded as irritating upon daily use.
A third group of mounting media is based on UV-curable polyacrylates. These offer the advantage that the mounted sample can be covered after a comparatively short exposure to UV irradiation with a coating of polyacrylate.
An example for a UV-curable mounting media is CureMount II (Electronic Microscopy Sciences, USA), which is based on isobornyl-acrylate as acrylate-monomer. However, this is a toxic monomer which exposes its users to health risks due to its volatility. The presentation “Energy Curable Oligomers for Outdoor Applications” (www.mycoatingresins.com Q2 2013) gives an overview on UV curable Oligomers.
US 3,467,617 discloses a UV curable mounting media which is non-fluorescent.
Ravikumar et al. in J. Dr. NTR University of Health Science, Vol. 3 (5), p. 1 -8, 2014 give a review on mounting media, inter alia also on UV curable mounting media.
EP 0104825 discloses in a general manner a curable adhesive composition, which has after curing a refractive index substantially similar to that of the sample. A preferred composition comprises a diacrylate-terminated polyurethane, a diacrylate ester of an epoxy resin and a diethoxyacetophenone initiator. It is formed on the mounting surface as a solution in a conventional organic solvent, such as isopropanol or toluene.
The UV curable mounting media of the prior art are usually a three component systems in which is difficult to regulate the viscosity. Is the mounting media to viscous it might form bubbles upon its application to the microscope slide which will make the subsequent microscopic analysis more difficult or even impossible. Further, the UV curable mounting media usually are based on monomers which are partial toxic and highly volatile.
Therefore, it was an object of the present invention to provide an UV curable mounting medium which has a suitable viscosity and which avoids the forming of bubbles upon mounting. Further, the amount of a Volatile Organic Compounds (VOC) should be small to not expose the user to volatile harmful substances.
A further requirement is the selective removal of the cover glass by an aprotic non-polar solvent to carry out a necessary staining or subsequent tests on the mounted sample.
This problem is solved by providing a UV curable mounting medium comprising a composition containing mono and/or bifunctional short chain acrylate oligomers or a mixture of mono- and a bifunctional short chain acrylate oligomer(s) with a molecular weight in the range from 100 to 1000 and a photopolymerisation initiator, wherein the photopolymerisation initiator is present in an amount of 0.81 to 3.5 wt%.
Surprisingly it was found that a cover glass which is mounted with the mounting medium according to the invention can be removed selectively after curing with xylene or toluene. In general the cover glass can be removed by an aprotic, non-polar solvent. Therefore, the composition according to the invention provides the easy possibility to allow additional staining or subsequent tests on the mounted tissue. Strong polar solvents, like acetone or acetic acid do not only remove a mounted sample from the cover glass but also from the mounting slide.
If the oligomer of the composition according to the present invention contains higher functional units, then a three-dimensional network is formed which cannot be removed by xylene, i.e. a removal of only the cover glass is impossible. If the amount of photopolymerisation initiator is below 0.89 wt%, the mounting medium does not harden within the standard hardening time of 30 to 120 seconds upon exposure to a UV/VIS light source, for example a commercial light source UV or LED (1 to 100 watt) with a maximum wavelength of 365 nm). Is the initiator present in an amount of more than 3.5 wt%, the cover glass cannot be remove with xylene at all even within 14 days. By using an amount of photopolymerisation initiator in the range of 1.95 to 3.5 wt%, the cover glass can be removed in a time span of more than 24 hours. The most preferred amount of photopolymerisation initiator is therefore in the range of 0.89, preferably 0.90 to 1.95 wt% which allows the selective removal of the cover glass after curing by a non-polar aprotic solvent within a commercially reasonable time, i.e. within 1 to 2 hours.
A further advantage of the mounting medium according to the present invention is the compatibility with washing solvents like alcohols as for example ethanol, isopropanol, and propanol, isoparaffines and petrol ether.
In the prior art, after staining of the sample, for example a tissue on the microscope slide with for example for hematoxyline, the tissue is usually washed with a sequence of several alcohols and before mounting washed with nonpolar non protic solvents to provide a compatibility with a mounting media and to remove the last rest of water.
In contrast to the mounting media in prior art, the present mounting media can also be applied directly on the samples or tissue cuts which are immersed in pure isopropanol or ethanol. This shortens the entire washing process and consumes less chemical solvents.
Further, the mounting medium according to the present invention shows a very good compatibility with xylene alternatives, like isoparaffines or petrol ethers or benzene fractions containing compounds of small chain lengths ( for example obtainable under the trademark Tissue Clear ® from Sakura). In a specific embodiment in so-called cover slipper machines an advantage of the use of the mounting medium according to the present invention is that the sample cannot dry in the cover slipper machine even upon staying a longer life span in the machine or during a malfunction when the tissue is washed before mounting with the mono and/or bifunctional acrylic oligomer or mixtures thereof.
A still further advantage of the mounting medium according to the invention is that a significantly lower amount of mounting medium can be used compared to physically curable mounting media in the prior art: only 50% of the amount of a mountain medium of the prior is necessary when using a mounting medium according to the invention to obtain the same effects and protection of the samples
In another preferred embodiment, the mounting medium has only at least one monofunctional short chain acrylate oligomers, i.e. monofunctional reactive units. Whereas in another also preferred embodiment the mounting media contains only at least bifunctional short chain acrylate oligomers, i.e. bifunctional units. In further specific embodiments, the mounting media comprises a mixture of at least one mono and at least one bifunctional short chain acrylate oligomers, i.e. mono and bifunctional reactive units.
The mounting medium according to the invention has preferably after curing a refractive index of 1.3 to 1.7. It is important that the refractory index of the mounting media is close to the refractory index of the cover glass and/or the mounting slide so that the resolution during analysis by a microscope is very good, to detect and determine very fine tissue structures etc. Is the refractory index higher or lower than the above mentioned limits, the resolution is markedly lower even if the contrast might be higher. If the mounting media has the same refractory index as the glass and is also in the range of the sample then, the sample would no longer be visible due to extinction of the wavelengths. As an indication only, the refractory index of a non stained histological sample is in the range of 1 .53 to 1.54 and a refractory index of a cover glass is around 1 .48 to 1.52 depending on the manufacturer Therefore, in a most preferred embodiment of the invention, a refractory index of the cured mounting medium is in the range of 1.47 to 1.55. Only for specific applications in microscopy (diatomaceous preparation, a refractory index of up to 1.7 is preferred (yellow medium). The lowest refractory index is 1.3 to obtain good results. Below this limit, no satisfactory resolution is possible.
In other embodiments of the present inventions provides a mounting medium comprises a mixture/composition of the following constituents:
A) 0 wt% to 80 wt% of an aliphatic urethane acrylate with a molecular weight of 800 to 2000; B) 20 wt% to 99 wt% of a short chain acrylate oligomer with a molecular weight from 150 to 1000 which is a mono and/or bifunctional short chain acrylate oligomer and
C) a photopolymerisation initiator,
Aliphatic urethane acrylates are all derivatives of urethanacrylates whose side chains consist of saturated or unsaturated branched or non-branched or cyclic aliphatic rests. Within the present invention, also the use of multifunctional urethane acrylates is possible, i. e. of urethane acrylate based compounds with more than one acryl group. Therefore, the aliphatic urethane acrylate according to the invention can comprise 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8 or at least 9 acryl groups. In especially preferred embodiments, the aliphatic urethane acrylate comprises two or more acryl groups. In a further especially preferred embodiment, the aliphatic urethane acrylate has exactly two acryl groups. The molecular weight of the aliphatic urethane acrylate according to the invention is in the range of 800 to 2000 g/mol, preferably between 1000 and 1500 g/mol and especially prefer between 1000 and 1200 g/mol.
As an aliphatic urethanacrylate, commercial available compounds can be used, for example Ebecryl 8402, Ebecryl 265, Ebecryl 837, Ebecryl 8254, Ebecryl 8409, Ebecryl 8465, Ebecryl 1291 , Ebecryl 1290 or Ebecryl 1259 (all available by Allnex, Belgium).
The mounting medium according to the invention comprises in an embodiment 40 wt% to 80 wt% and to 80 wt%, preferably 50 wt% to 70 wt%, most preferred 55 wt% to 65 wt% in specific embodiments 60 wt%, 63 wt% or 65 wt% of the aliphatic urethane acrylate. Is the amount of the aliphatic urethane acrylate to low, that is especially lower than 40 wt%, then, the mounting medium according to the invention is less resistant to pressure after curing and may crack.
Further, the mounting media according to the invention comprises a short chain acrylate oligomer. The term“short chain acrylate oligomer” in the sense of the present invention means a acrylate oligomer with a chain length of 2 to 9 acrylic acid units (monomers) wherein basically all chain length, i. e. chains comprising of 2, 3, 4, 5, 6, 7, 8 and 9 monomers are suitable for carrying out the present invention especially preferred oligomers consisting of 3 to 8 monomers. The side chains of the short chain acrylate oligomers can also be modified saturated or unsaturated, branched or unbranched or cyclic aliphatic rests. Within the scope of the present invention, also the use of multifunctional short chain acrylate oligomers is contemplated, i. e. acrylate compounds of more than one acryl group. In a further preferred embodiment, the short chain acrylate oligomer comprises three or more acryl groups (monomers). In a further embodiment, the short chain acrylate oligomer comprises exactly three acryl groups (monomers). The term also comprises molecules with only one acryl group linked to a monomeric aliphatic or olefin containing chain or a“spacer” chain between two acrylic groups, wherein the chain and the“spacer” chain are modified saturated or unsaturated, branched or unbranched or cyclic aliphatic rests.
The molecular weight of the short chain acrylate oligomer according to the invention is between 150 to 1000 g/mol, preferably between 300 und 800 g/mol, an most referred between 450 to 600 g/mol.
As short chained acrylate oligomer commercially available compounds can be used, for example Ebecryl LEO 10501 , Ebecryl 130, Ebecryl 140, Ebecryl 150 or Ebecryl 160, Ebecryl 1 13, Ebecryl 114, Ebecryl 11 or HDDA , especially preferred are Ebecryl 1 13, Ebecryl 114, Ebecryl 11 and HDDA (1 ,6-hexanediol diacrylate) (all available from Allnex, Belgium).
The mounting medium according to the invention comprises 20 wt% to 60 wt%, preferably 30 wt% to 50 wt%, most preferred 25 wt%, 30 wt%, 35 wt% or 40 wt% of the short chain acrylate. Is the amount of aliphatic urethane acrylate too low, the viscosity of the mounting media according to the invention is too high which is difficult in handling and prevents its use in cover slipper machines.
In one embodiment, the aliphatic urethane acrylate and/or the short chain acrylate oligomer are multifunctional. In a preferred embodiment, both components are multifunctional. In a further preferred embodiment, the aliphatic urethane acrylate is bifunctional or trifunctional and the short chain acrylate oligomer is biifunctional. By adding additional acryl groups, the polymerization characteristics of the mounting medium according to the invention can be influence, since crosslinking may occur.
As a photopolymerisation initiator, all compounds can be used in the present invention which form during UV irradiation radicals. For example, photopolymerisation initiators from the group of acetophenones, benzyl or benzoine compounds, benzophenones, cationic photopolymerisation initiators or thioxanthone alone or in combination with one another can be used.
As a photopolymerisation initiator, commercially available compounds can be used alone or in combination thereof, for example Omnirad 184, Omnirad 500, Omnirad 1173, Omnirad 2959, Omnirad 819, Omnirad MBF or Omnirad TPO (all available from IGM Resins, Netherlands). Especially preferred is Omnirad 1 173 alone. IGM Resins has taken over production of photopolymerisation initiators from BASF with former Tradename Irgacure.
In a further embodiment, the mounting medium according to the invention comprises 0.5 to 3.5 wt% of the photopolymerisation initiator. By optimizing the amount of photopolymerisation initiator the polymerisation time and the storage can be advantageously also the polymerisation before application can be avoided. In a specific preferred embodiment, the amounting medium according to the invention comprises 0.9 to 1.95 wt%, especially 1.0 to 3.5 wt%, especially 1.0 to 1.95 wt% of the photopolymerisation initiator as already discussed above.
In further preferred embodiments the mounting medium according to the invention comprises ca. 0.1 wt%, 1.0 wt%, 1.2 wt%, 1.3 wt%, 1.4 wt%, 1.5 wt%, 1.6 wt%, 1 .7 wt%, 1.8 wt% of the photopolymerisation initiator. In a further preferred embodiment, the photopolymerisation initiator of the mounting medium according to the invention is a short or long wave photopolymerisation initiator or a mixture thereof. By the use of photopolymerisation initiators which react under exposure to UV light of different wave length the mounting medium according to the invention can be adapted to different areas of use and UV sources. By the use of a mixture of photopolymerisation initiators which react on exposure to UV light of different wave lengths, the mounting medium according to the invention can be polymerized with a multitude of various UV sources.
The viscosity of mounting media can be measured by different tools. In case of UV- curable mounting media a flow cap-system is used (according DIN 53 21 1 ). The ideal viscosity for a classic system like EUKITT® (physically drying) is around 120 sec flow time in DIN cap 4 (at 20°C). For UV-curable systems the viscosity has to be lower because the time to penetrate the tissue is shorter as the time for the curing-process is much faster. The ideal viscosity range of UV-curable mounting media according to the present invention (especially when used in cover slipping machines) is between 15 seconds and 90 seconds in a DIN 4 flow cap system at 20°C. The mounting medium according to the invention has a preferred viscosity in the range of 20 to 70 sec flow time in DIN cap 4 (at . 20°C), still more preferred 40-60 and in further specific embodiments 45, 50 and 55 sec flow time in DIN cap 4 (at 20°C). This provides in one aspect an easier handling and in another aspect the formation of bubbles during mounting is avoided. The mounting medium according to the invention is easy to store and can also be used by applying usual security measures in chemical laboratories.
The mounting medium according to the invention contains only a small amount of highly volatile toxic compounds and avoids therefore the application of specific security provisions. Upon irradiation with UV light, it is hardening very fast and without a shrinking. After hardening, it is completely transparent and can be stored over a long period of 1 to 15 years without discolor or forming cracks Due to its viscosity it is also advantageously useable for cover slipper machines. It is possible to obtain a mounted sample without forming bubbles and the mounting medium according to the invention conserve the samples without altering its structure or color. Further, the mounted samples are protected against humidity, oxygen, day light and UV light and further external influences and protect the samples against degradation. Further, the mounting medium according to the invention has positive polarization properties and shows no polarization colors or lines on mounting slides and cover glasses. The term absorption maximum (UV/VIS) of a substance is its absorption maximum in the range of UV and visible light. The absorption maximum (UV/VIS) of a substance determines its color. Transparent substances have an absorption maximum (UV/VIS) of less than 380 nm, yellow substances an absorption maximum of 440 nm. Substances which do not alter its absorption maximum substantially even after several years do not alter its color and do not show a yellowing. A non-substantial change of absorption maximum is present if the absorption maximum changes within three years less than 30 nm. In a further specific embodiment, the mounting medium according to the invention is characterized in that it is colorless after hardening/curing and the difference in the absorption maximum measured after one day after hardening/curing and after exactly three years after hardening/curing is less than 30 nm.
Therefore, the mounting medium according to the invention has even after a long storage no or only a small staining effect and is therefore also extremely suitable for the mounting of samples which have to be examined over a long period of time on several occasions. A mounting medium has also to have a specific refractive index after curing. This refractive index is to be adapted to the mounted tissue as also to the microscope objective with which the sample is examined. The refractory index of the mounting medium can be influenced by its chemical composition. Especially by mixing different chemical compounds, a desired refractory index can be obtained. The person skilled in the art is well aware of methods for the measuring of the indetermination of the refractory index, for example by use of a refractometer. In specific embodiments of the present invention the refractory index of the mounting medium is after curing in the range from 1.3 to 1.7.
The mounting medium of the present invention can be manufactured by processes known essentially to a person skilled in the art. For example, the mounting medium according to the invention can be obtained by mixing of the separate components. Either all of the components can be mixed together or two components are mixed in a first step and all the other components can be added in one or more steps.
Still further embodiment the composition for the mounting medium can be used as a nail polish or as a glue. The mounting medium of the present invention can comprise further components for example inert solvents, diluents, indicators, light stabilizers, antioxidants, pigments or colors.
The mounting medium according to the invention can be used as any other mounting medium known in the art. Biological samples which have to be mounted with the mounting medium according to the invention comprise cells, bacteria, tissue cuts, tissue samples, blood, plasma or serum samples. The samples can either be directly mounted or previously stained. The staining methods are known essentially known to a person skilled in the art and comprise for example hematoxyline-eosin staining, Giemsa staining, gram staining and toluidine staining. The mounting medium according to the invention can be used for the mounting of flat samples liker cells or tissue cuts or mounting of three dimensional structures as tissues, insects, plants or pieces thereof. Further, the mounting medium according to the invention is especially suitable for the mounting of samples with low contrast or colorless samples.
The mounting medium according to the invention can be hardened/cured by any conventional UV source, for example with UV lamps or UV hardening devices. The irradiation time depends on the exact composition of the mounting medium according to the invention. A person skilled in the art is aware of how to determine the optimum irradiation time and intensity. Preferably, UV lights source with a wave length range of 220 to 360 nm during 120 sec. are used for curing at the mounting medium according to the invention.
The mounting medium according to the invention hardened after applying UV irradiation within 120 sec., preferably within 30 sec.
The mounting medium according to the invention is also suitable for the mounting of fluorescing and fluorescing stained samples. In a further embodiment the mounting medium according to the invention contains an anti-bleaching agent like BHT (Butylhydroxytoluol), Irganox 1076 (Octadecyl 3-(3,5-di-tert-butyl-4-hydroxyphenyl) propionate), Irganox 1010 (Pentaerythrittetrakis (3-(3,5-di-tert-butyl-4-hydroxyphenyl) propionate), Tinuvin 312 (N-(2-Ethoxyphenyl)-N’-(4-thylphenyl)-ethlyene diamid), Tinuvin 1577 (2-(4,6-diphenyl-1 ,3,5-triazin-2-yl)-5-(hexyl)oxyphenol) and other suitable agents alone or in combination thereof which avoid a bleaching of the fluorescence staining agent in the sample. The UV curable mounting medium according to the invention can also be used on cover slipping films (like cellulose triacetate films - or TAC-films) which are widely used in cover
slipping machines/machines.
The present invention is further explained by the following Examples which should not be
understood as limiting the scope of the present invention. Exa
Example 1
180.0 g (98.435 wt%) Ebecryl 113 were placed in a 500 ml flask, and under mixing (IKA
Eurostar 200 P4 Control), Irgacure 1 173 (2.862 g (1.565 wt%) is mixed with a dissolver
disk (diameter 6 cm). The mixture was then mixed during 15 minutes at room temperature and bottled and
stored in an aluminum bottle.
Example 2
182.0 g (99.10 wt%) Ebecryl 113 were placed in a 500 ml flask, and under mixing (IKA
Eurostar 200 P4 Control), Irgacure 1173 (1 .653 g (0.90 wt%) was mixed with a dissolver
disk (diameter 6 cm) and with a IKA RW 27 D mixer.
The mixture was then mixed during 15 minutes at room temperature and bottled and
stored in an aluminum bottle.
Example 3
195.0 g (98.784 wt%) Ebecryl 114 were placed in a 500 ml flask, and under mixing (IKA
Eurostar 200 P4 Control), Irgacure 1 173 (2.40 g (1.216 wt%) was mixed with a dissolver
disk (diameter 6 cm) and with a IKA RW 27 D mixer.
The mixture was then mixed during 15 minutes at room temperature and bottled and
stored in an aluminum bottle. Example 4
225.0 g (98.05 wt%) Ebecryl 1 1 were placed in a 500 ml flask, and under mixing (IKA Eurostar 200 P4 Control), Irgacure 1173 (4.475 g (1.95 wt%) was mixed with a dissolver disk (diameter 6 cm) and with a IKA RW 27 D mixer. The mixture was then mixed during 15 minutes at room temperature and bottled and stored in an aluminum bottle.
Example 5
185.0 g (98.25 wt%) HDDA were placed in a 500 ml flask, and under mixing (IKA Eurostar 200 P4 Control), Irgacure 1 173 (3.238 g (1.75 wt%) was mixed with a dissolver disk (diameter 6 cm) and with a IKA RW 27 D mixer.
The mixture wass then mixed during 15 minutes at room temperature and bottled and stored in an aluminium bottle.
All mounting media of examples one to five were cured within 30 to 120 seconds with a commercially available light source (UV or rather LED 1 - 100 Watt) at a wavelengths maximum of 365 nm. A cover slipped mounted on a mounting slide was selectively removed with xylene within 24 hours.
Example 6
168.6 g (61.16 wt%) Ebecryl 8402 were placed in a 500 ml flask and heated in a water bath at 60° C by mixing (IKA Eurostar 200 P4 Control) with a dissolver disk (diameter 6 cm). In a second 250 ml flask, 4.24 (1.53 wt%) Irgacure 1 173 in 102.85 g (37.31 wt%) Ebecryl LEO 10501 were dissolved and mixed and heated in a water bath at 60° C with a dissolver disk (diameter 3 cm) and with a IKA RW 27 D mixer.
The Irgacure 1 173 which was completely dissolved in Ebecryl LEO 10501 was added to the preheated Ebecryl 8402. The mixture was mixed for further 15 minutes and bottled in an aluminum bottle. Example 7
180.3 g (67.35 wt%) Ebecryl 1259 were placed in a 500 ml flask and heated in a water bath at 60° C by mixing (IKA Eurostar 200 P4 Control) and mixed with a dissolver disk (diameter 6 cm). In a second 250 ml flask, 5.0 (1.87 wt%) Darocur TPO and 2.0 g (0.75 wt%) Darocur MBF in 80.4 g g (30.03 wt%) Ebecryl LEO 10501 was dissolved under mixing and heating in water bath at 60° C with a dissolver disk (diameter 3 cm) and a IKA RW 27 D mixer
The Darocur TPO/MBF which was completely dissolved in Ebecryl LEO 10501 was added to the preheated Ebecryl 1259. The mixture was mixed for further 15 minutes and bottled and stored in an aluminum bottle.
Example 8
148.23 g (42.35 wt%) Ebecryl 265 were placed in a 500 ml flask and heated in a water bath at 60° C by mixing (IKA Eurostar 200 P4 Control) and mixed with a dissolver disk (diameter 6 cm). In a second 250 ml flask, 8.75 (2.5 wt%) Irgacure 184 in 139.48 g (39.85 wt%) Ebecryl LEO 140 was dissolved by mixing and heating in water bath at 60° C with a dissolver plate (diameter 3 cm) and IKA RW 27 D mixer.
The Irgacure 184 which was completely dissolved in Ebecryl 140 was added to the preheated Ebecryl 265. The mixture was mixed for further 15 minutes and bottled in an aluminum bottle. Example 9
160 g (44.0 wt%) Ebecryl 265 were placed in a 500 ml flask and heated in a water bath at 60° C by mixing (IKA Eurostar 200 P4 Control) and mixed with a dissolver disk (diameter 6 cm). In a second 250 ml flask, 6.55 (1.8 wt%) Irgacure 819 in 197.1 g (54.2 wt%) Ebecryl 160 was dissolved under mixing and heating in water bath at 60° C with a dissolver disk (diameter 3 cm) and IKA RW 27 D mixer.
The Irgacure 819 which was completely dissolved in Ebecryl 160 was added to the preheated Ebecryl 265. The mixture was mixed for further 15 minutes and bottled and stored in an aluminum bottle.

Claims

Patent Claims
1. UV curable mounting medium comprising a composition containing a short chain acrylate oligomer with one or two reactive functional groups or a mixture thereof with a molecular weight of 100 to 1000;
a photopolymerisation initiator in an amount of 0.89 to 3.5 wt%.
2. UV curable mounting medium according to claim 1 wherein the amount of the photopolymerisation initiator is in a range of 0.89 to 1.95 wt%.
3. UV curable mounting medium according to claim 1 or 2, wherein the short chain acrylate oligomer has two functional groups.
4. UV curable mounting medium according to one of the preceding claims comprising
A) 40 wt% to 80 wt% of an aliphatic urethane acrylate within a molecular weight of 800 to 2000;
B) 20 wt% to 60 wt% of a short chain acrylate oligomer with a molecular weight of 150 to 1000;
C) a photopolymerisation initiator in an amount of 0.89 to 3.5 wt%.
5. Mounting medium according to claim 4 wherein components A) and/or B) are multifunctional acrylates.
6. Mounting medium according to claim 5 wherein component A) has two functional groups.
6. Mounting medium according to one of the preceding claims wherein the photopolymerisation initiator is a short- or longwave photopolymerisation initiator or a mixture thereof.
7. Mounting medium according to one of the preceding claims, wherein the mounting medium is after curing colorless and wherein the difference of the absorption maxima measured at day one and three years after curing is smaller than 30 nm.
8. Mounting medium according to one of the preceding claims 4 to 8 comprising 63 wt% of component A) and 35 wt% of component B).
9. Use of a mounting medium according to one of the preceding claims on cover slipping films in cover slipper machines.
PCT/EP2017/001407 2017-12-06 2017-12-06 Uv curable mounting medium WO2019110078A1 (en)

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