WO1996031549A1 - Polymeres greffes dendrimeres - Google Patents

Polymeres greffes dendrimeres Download PDF

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Publication number
WO1996031549A1
WO1996031549A1 PCT/EP1995/001278 EP9501278W WO9631549A1 WO 1996031549 A1 WO1996031549 A1 WO 1996031549A1 EP 9501278 W EP9501278 W EP 9501278W WO 9631549 A1 WO9631549 A1 WO 9631549A1
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WIPO (PCT)
Prior art keywords
dendrimeric
separation
formula
radical
graft polymers
Prior art date
Application number
PCT/EP1995/001278
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German (de)
English (en)
Inventor
Egbert Müller
Margot Mack
Peter Poguntke
Dieter Lubda
Original Assignee
Mueller Egbert
Margot Mack
Peter Poguntke
Dieter Lubda
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 Mueller Egbert, Margot Mack, Peter Poguntke, Dieter Lubda filed Critical Mueller Egbert
Priority to PCT/EP1995/001278 priority Critical patent/WO1996031549A1/fr
Publication of WO1996031549A1 publication Critical patent/WO1996031549A1/fr

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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/281—Sorbents specially adapted for preparative, analytical or investigative chromatography
    • B01J20/286—Phases chemically bonded to a substrate, e.g. to silica or to polymers
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/30—Processes for preparing, regenerating, or reactivating
    • B01J20/32—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating
    • B01J20/3202—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating characterised by the carrier, support or substrate used for impregnation or coating
    • B01J20/3204—Inorganic carriers, supports or substrates
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/30—Processes for preparing, regenerating, or reactivating
    • B01J20/32—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating
    • B01J20/3202—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating characterised by the carrier, support or substrate used for impregnation or coating
    • B01J20/3206—Organic carriers, supports or substrates
    • B01J20/3208—Polymeric carriers, supports or substrates
    • B01J20/321—Polymeric carriers, supports or substrates consisting of a polymer obtained by reactions involving only carbon to carbon unsaturated bonds
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/30—Processes for preparing, regenerating, or reactivating
    • B01J20/32—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating
    • B01J20/3231—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating characterised by the coating or impregnating layer
    • B01J20/3242—Layers with a functional group, e.g. an affinity material, a ligand, a reactant or a complexing group
    • B01J20/3268—Macromolecular compounds
    • B01J20/327—Polymers obtained by reactions involving only carbon to carbon unsaturated bonds
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/30—Processes for preparing, regenerating, or reactivating
    • B01J20/32—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating
    • B01J20/3231—Impregnating or coating ; Solid sorbent compositions obtained from processes involving impregnating or coating characterised by the coating or impregnating layer
    • B01J20/3242—Layers with a functional group, e.g. an affinity material, a ligand, a reactant or a complexing group
    • B01J20/3268—Macromolecular compounds
    • B01J20/3278—Polymers being grafted on the carrier
    • 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
    • C08F285/00—Macromolecular compounds obtained by polymerising monomers on to preformed graft polymers
    • 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
    • C08F291/00—Macromolecular compounds obtained by polymerising monomers on to macromolecular compounds according to more than one of the groups C08F251/00 - C08F289/00
    • C08F291/06—Macromolecular compounds obtained by polymerising monomers on to macromolecular compounds according to more than one of the groups C08F251/00 - C08F289/00 on to oxygen-containing macromolecules
    • C08F291/08—Macromolecular compounds obtained by polymerising monomers on to macromolecular compounds according to more than one of the groups C08F251/00 - C08F289/00 on to oxygen-containing macromolecules on to macromolecules containing hydroxy radicals
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2220/00—Aspects relating to sorbent materials
    • B01J2220/50—Aspects relating to the use of sorbent or filter aid materials
    • B01J2220/54—Sorbents specially adapted for analytical or investigative chromatography

Definitions

  • the invention relates to dendrimeric graft polymers and their use as separation materials for liquid core chromatography.
  • separation effectors required for the various chromatographic separation processes are bound as a solid phase to a base support and interact with the analytes of the sample to different extents.
  • Suitable separation effectors are known to the person skilled in the art for various chromatographic separation methods; for example: ionic or ion-forming (ionogenic) groups for ion exchange chromatography, affinity ligands for affinity chromatography, which also includes metal chelate chromatography, hydrophobic groups for hydrophobic interaction chromatography or reversed phase chromatography and predominantly reticulated porous hydrophilic groups for gel permeation chromatography.
  • the invention relates to dendrimeric graft polymers based on hydroxyl-containing base supports, on the surface of which polymers are covalently bonded, with a) the base support containing aliphatic hydroxyl groups, b) the covalently bonded polymers being bonded to the base support via a terminal monomer unit, c) the polymers contain monomer units of the formula II at the branching points of the dendrimeric structure
  • d-Cj-alkyl or C 6 -C 12 aryl, n represents an integer between 1 and 5, one radical X OH and the other radical X represents a terminal monomer unit of a further polymer chain, and
  • Y is a radical containing a separation effector.
  • the dendrimeric graft polymers according to the invention can be obtained by the following reaction steps: a) grafting of monomers of the formula I onto a hydroxyl group-containing base support in the presence of cerium IV ions,
  • Ri, R2, R3, R and n have the meanings already mentioned; b) at least partially converting the oxirane groups into diol groups; c) grafting on further monomers, for example further monomers of the formula I, or else monomers which contain separation effectors in the presence of cerium IV ions; d) optional repetition of steps b) and c); e) introduction of residues with separation effectors, insofar as these have not already been introduced by the grafting reaction with monomers which contain separation effectors; f) optional ring opening of remaining oxirane groups.
  • the invention relates to the use of the dendrimeric graft polymers according to the invention in the separation of mixtures of at least two substances, in particular for the separation of biopolymers, by means of liquid chromatography, in particular by means of ion exchange and affinity chromatography.
  • Another use according to the invention is the separation of low molecular weight analytes from protein-containing matrices.
  • the invention also relates to processes for the preparation of dimeric graft polymers with the following process steps: a) grafting of monomers of the formula I onto a hydroxyl group-containing base support in the presence of cerium IV ions,
  • RH, C r C 5 alkyl or C 6 -C 12 aryl and n are an integer between 1 and 5, with activated carrier materials known from DE 43 10 964 being formed, b) at least partially converting the oxirane groups into diol groups; c) grafting of monomers of the formula I or of monomers of the formula V onto the diol groups formed in step b) in the presence of cerium (IV) ions,
  • R 8 C -C 0 alkyl which with an amino, monoalkylamino
  • Dialkylamino, trialkylammonium, carboxyl or AC sulfonic acid residue is substituted, where steps b) and c) can be repeated one or more times, • d) introduction of the residues which contain the separation effectors, insofar as Q has not already done so the grafting reaction with monomers of
  • the invention relates to processes for the separation of mixtures of at least two substances, in particular for the separation of biopolymers, by means of liquid chromatography, in particular by means of ion exchange, hydrophobic interaction or affinity chromatography, using the dendrimeric graft polymers according to the invention. Further methods according to the invention relate to the separation of low molecular weight analytes from protein-containing matrices with the aid of dendrimeric separation materials according to the invention.
  • Figure 1 shows the dependence of selectivity ⁇ (curve A) and binding capacity (curve B) on diethylamine-substituted (DEA) ion exchangers with different degrees of branching (sample number as abscissa).
  • curve A selectivity
  • curve B binding capacity
  • DEA diethylamine-substituted
  • Figures 2 and 3 show the experimental setup for the separation of analytes from biological matrices, for example serum or plasma, using separation materials according to the invention.
  • Figure 4 shows the results of a recovery test using a separating material produced according to Example 9. Further experimental details can be found in application example B.
  • the dendrimeric graft polymers according to the invention are distinguished by a tree-like branched structure, a first linear polymer being grafted onto a base support which contains aliphatic hydroxyl groups.
  • Monomers of the formula I, where the radicals R 1 , R2, R3 and R 4 and n have the meanings already mentioned, are grafted on in the presence of cerium IV ions, a linear graft polymer being formed.
  • the basics of this reaction are described by G. Mino and S. Kaizerman (1958) J. Polymer Science __1, 242-243, and G. Mino et al. (1959) J. Polymer Science __, 393-401.
  • the polymer initially contains oxirane residues, which are then completely or partially converted into diol groups. Treatment with dilute sulfuric acid is preferred.
  • Monomers of the formula I can now be polymerized again in the presence of cerium IV ions onto the aliphatic hydroxyl groups of this polymer (first generation) thus formed.
  • the resulting polymer (second generation) is linear in itself. However, the forest is branched.
  • the above-mentioned steps can be repeated: conversion of the oxirane groups into diol groups and polymerization of monomers of the formula I in the presence of cerium IV ions.
  • separation effectors required for the chromatographic separations are introduced, which are bound to a base support as a solid phase and which interact to different degrees with the analytes of the sample.
  • Suitable separation effectors are known to the person skilled in the art for various chromatographic separation methods, for example: a) For ion exchange chromatography, ionic groups such as, for example, quaternary ammonium alkyl groups and the SO 3 group, and also ionogenic groups which form ions under certain pH conditions are known. The last group includes, for example, the alkylated amino groups, as well as the carboxyl and phosphoric acid groups. b) A large number of affinity ligands are known to those skilled in the art for affinity chromatography, each of which forms a structurally defined bond with the analyte, and which as
  • Separation effectors are suitable, for example:
  • Affinity ligand analyte (example.
  • Uncharged hydrophobic separation effectors are customary for hydrophobic interaction chromatography, for example C 1 -C 2 o-alkyl,
  • hydrophilic compounds which preferably form pores or networks, are used as the separation effector.
  • These include (meth) acrylic acid derivatives such as acrylamide or methacrylamide, furthermore (2,3-dihydroxypropyl) methacrylate or N- (2-methoxyethyl) acrylamide or N- (2,3-dihydroxypropyl) acrylamide.
  • They also include vinylated heterocycles such as 1-vinylimidazole, N-vinylpyrrolidone, 2-vinylpyridine, 4-vinylpyridine, 4-vinylpyrrolidone-N-oxide.
  • So-called shielded phases are used for the separation of low molecular weight substances from biological samples (eg urine or blood). These separating materials have both hydrophobic and hydrophilic areas.
  • the hydrophobic areas the structure of which corresponds to the hydrophobic separation materials mentioned above (see c)), interact with the low molecular weight analyte of the sample.
  • the hydrophilic areas prevent the interaction of the high molecular weight portions of the sample (eg the proteins) with the hydrophobic areas.
  • Dendrimeric graft polymers according to the present invention are outstandingly suitable as shielded phases for the
  • Oxirane residues which are present after the polymerization with compounds of the formula I are built into the support; e.g .: a1) the reaction with sulphurous acid or its salts or with primary, secondary or tertiary amines, whereby ion exchangers are formed; a2) the reaction with iminodiacetic acid or the introduction of thiophilic ligands, or other affinity ligands such as protein A, producing carriers for affinity chromatography; a3) the reaction with alcohols, phenols or primary amines, whereby hydrophobic separation materials are formed.
  • Hydrophobic separation effectors can also be introduced, for example, by ester bonds to hydroxyl groups, such as those formed by hydrolysis of the oxirane residues.
  • R 4 is H, d-Cj-alkyl or C 6 -C 12 aryl, n is an integer between 1 and 5, one radical Z OH and the other radical Z is a radical selected from the group NR5R6, N + R5R6R ?, po 4 H 2 and S0 3 H, and Rs, R6 and R 7 independently
  • R 8 CrCio-alkyl which is substituted with an amino, monoalkylamino, dialkylamino, trialkylammonium, carboxyl or sulfonic acid residue.
  • radical Y from formula III denotes a radical according to formula VI
  • Preferred monomers of the formula V are those in which W has one of the following meanings: OH, NH (CH 2 ) 2 N (CH 3 ) 2 , NH (CH 2 ) 2 N (C 2 H 5 ) 2 , NH (CH 2 ) 2 N + (CH 3 ) 3 , NHC (CH 3 ) 2 CH 2 S0 3 H or NH (CH 2 ) 2 S0 3 H.
  • any remaining oxirane residues can be hydrolyzed, for example by a final treatment with dilute sulfuric acid.
  • room temperature (RT) means
  • the polymerization is carried out in a suitably sized three-necked flask equipped with a stirrer, dropping funnel and thermometer. It is washed by suction on a glass frit (G2).
  • Example 1 Production of an oxirane-activated carrier starting from Fractogel®-TSK HW 65 (S)
  • a suspension of 100 ml of sedimented Fractogel®-TSK HW 65 (S) and 66 ml of water is mixed with 3 g of ammonium cerium (IV) nitrate (dissolved in a
  • Example 2 Production of an oxirane-activated carrier starting from LiChrospher® diol
  • Example 3 Grafting a polymer onto a diol-containing polymer (producing the dendrimeric structure)
  • Stage 1 Transfer of the oxirane into diol groups 100 ml of extracted oxirane-activated carrier material produced according to
  • Example 1 are hydrolyzed with 200 ml of 0.5 M sulfuric acid (1 hour, 50 ° C.) and the oxirane groups are thus converted into diol groups. It is then washed three times with 200 ml of water each time.
  • Step 2 grafting on another polymer chain
  • the result is a single-branched dendrimary material with oxirane residues.
  • Example 4 Grafting a polymer onto a diol-containing polymer (generation of multi-branched denim structures)
  • Example 3 The material from Example 3 is again subjected to the reaction sequence described in Example 3. A doubly branched denim material with oxirane residues is formed.
  • the washed reaction product is suspended in 100 ml of a 0.5 M sulfuric acid solution and slowly stirred at 40 ° C. for two hours. Then it is washed with 0.25 M phosphate buffer (pH 7) to the neutral point, then with water. The gel is stored in aqueous suspension with the addition of 0.02% sodium azide.
  • a solution of 15 g NaOH and 25 g iminodiacetic acid in 100 ml water is adjusted to pH 11 with concentrated HCl and decolorized with 1 g activated carbon.
  • 50 ml of suctioned-off oxirane-activated carrier material prepared according to Example 4 (branched twice) are added to this solution.
  • the solution is stirred at 45 ° C for 20 hours.
  • the reaction product is filtered off, washed with 250 ml of 0.5 M NaOH and washed with water.
  • the unreacted oxirane groups were hydrolyzed by treatment with 100 ml of 0.5 M sulfuric acid (2 hours, 45 ° C.).
  • the affinity carrier material is then washed once with 100 ml of 0.5 M sulfuric acid, twice with 100 ml of water, once with 100 ml of 0.5 M phosphate buffer pH 7 and once with 100 ml of 1 M NaCl solution.
  • the gel is stored in 0.02 M phosphate buffer pH 7 with the addition of 1 M NaCl and 0.02% NaN 3 .
  • Step 1
  • Step 1 Preparation of a Glycidyloxypropyl Carrier
  • Stage 2 Ring opening of the glycidyloxypropyl carrier to the diol phase.
  • the product obtained from stage 1 is suspended in 50 ml of a 5% sulfuric acid solution and refluxed for 3 hours with slow stirring to open the epoxy ring. After the reaction suspension has been suctioned off, it is washed free of sulfate with water and, after washing again, is dried with methanol.
  • a diol carrier (carbon content 7.6%; corresponding to 2.81 mmol / m ⁇ diol groups) is obtained.
  • Stage 2 and 60 ml of water are mixed with 0.8 g of ammonium cerium (IV) nitrate (dissolved in a mixture of 40 ml of water and 1.2 g of HNO 3 (65%)) at room temperature with vigorous stirring. After 1 minute, a solution of 1.2 g (2,3-epoxypropyl) methacrylate in 15 ml of dioxane is added. It is stirred for an hour. The reaction product is then washed twice with 200 ml of water, three times with 100 ml of acetone and three times with 200 ml of water. Stage 4: sulfuric acid hydrolysis
  • Step 5 Second graft polymerization
  • Step 6 Reaction of the dendrimeric graft polymer with hydrophobic
  • Ligands 5 g of the dried carrier material from stage 5 are suspended at 0 ° C. in dry chloroform. 60 ml of dried triethylamine are added to this solution. A solution of 2 g of stearoyl chloride in 25 ml of chloroform is then added over the course of 3 hours, while cooling to 4 ° C.
  • the mixture is stirred at room temperature for 48 hours and the gel is then washed with 50 ml of chloroform, methanol, water and methanol and dried.
  • a dendrimeric shielded phase separation material is provided, the hydrophobic residues of which are shielded by hydrophilic diol groups.
  • the following application example shows the influence of the degree of branching on the binding capacity and separating capacity of the separating material.
  • Example 2-7 Single to seven-fold branched DEA-derived separation materials (samples 2-7) are produced in accordance with examples 5 and 6, unbranched comparison material (sample 1) is produced in analogy to example 5, the linear polymer from example 1 being used instead of the dendrimeric oxirane-containing polymer becomes.
  • Waste container (devices from E. Merck, Darmstadt, Germany)
  • Fig. 3a Position "L” (LOAD)
  • Fig. 3b Position "I” (INJECT)
  • Guard column buffer (1) 0.05 M Na phosphate (pH 5.0); analytical column ((8);
  • the sample After the plasma sample (100 ⁇ l) has been injected by the automatic sample dispenser (5) into position "L" of the changeover valve (6), the sample reaches the pre-column buffer ((1); pumped by the HPLC pump (3); flow rate 0 , 5 ml / min) on the guard column (7).
  • the analyte (carbamazepine) is selectively retained by the separating material of the guard column, while matrix components, in particular proteins, are eluted into the waste container (11) within 12 minutes.
  • the elution diagrams for A are: a calibrator containing 0.5 ⁇ g carbamazepine, and
  • B a plasma sample (human plasma) which also contains 0.5 ⁇ g carbamazepine.
  • the analyte is completely found again in the plasma sample.

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  • Organic Chemistry (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Analytical Chemistry (AREA)
  • Health & Medical Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Polymers & Plastics (AREA)
  • Inorganic Chemistry (AREA)
  • Treatment Of Liquids With Adsorbents In General (AREA)
  • Solid-Sorbent Or Filter-Aiding Compositions (AREA)

Abstract

L'invention concerne des polymères greffés dendrimères comprenant des supports de base porteurs de groupes hydroxyle et sur la surface desquels des polymères sont liés par covalence, caractérisés en ce que a) les supports de base renferment des groupes hydroxyle aliphatiques, b) les polymères liés par covalence sont liés aux supports de base par l'intermédiaire d'un motif monomère terminal, c) les polymères renferment, aux points de ramification de la structure dendrimère, des motifs monomères de formule (II), d) les motifs monomères des polymères dendrimères renferment un composé de formule (III) dans laquelle R?1, R2 et R3¿ désignent, indépendamment l'un de l'autre, H ou CH¿3, R?4 désigne H, un alkyle en C¿1?-C5 ou un aryle en C6-C12, n est un nombre entier compris entre 1 et 5, un reste X est un OH, et l'autre X représente un motif monomère terminal d'une autre chaîne polymère, et Y désigne un reste renfermant un activateur de séparation. L'invention concerne en outre la fabrication de ces polymères greffés dendrimères et leur application comme matériaux séparateurs en chromatographie en phase liquide.
PCT/EP1995/001278 1995-04-07 1995-04-07 Polymeres greffes dendrimeres WO1996031549A1 (fr)

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Cited By (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1998040502A1 (fr) 1997-03-14 1998-09-17 Life Technologies, Inc. Transfections activees par des peptides
WO2004003542A1 (fr) * 2002-06-28 2004-01-08 Amersham Biosciences Ab Matrices de base a surface modifiee
WO2014067605A1 (fr) * 2012-11-01 2014-05-08 Merck Patent Gmbh Modification superficielle de supports de base poreux
US9303098B2 (en) 2008-05-30 2016-04-05 Merck Patent Gmbh Ce(IV)-initiated graft polymerisation on polymers containing no hydroxyl groups
US9358300B2 (en) 1998-11-12 2016-06-07 Life Technologies Corporation Transfection reagents
US10195280B2 (en) 2014-07-15 2019-02-05 Life Technologies Corporation Compositions and methods for efficient delivery of molecules to cells
EP3460065A1 (fr) 2012-04-20 2019-03-27 Commonwealth Scientific and Industrial Research Organisation Procédé de transfection cellulaire
US11606940B2 (en) 2015-08-07 2023-03-21 Commonwealth Scientific And Industrial Research Organisation Method for producing an animal comprising a germline genetic modification

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1994026379A1 (fr) * 1993-05-13 1994-11-24 Merck Patent Gmbh Procede et entraineurs pour la chromatographie par permeation de gel

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1994026379A1 (fr) * 1993-05-13 1994-11-24 Merck Patent Gmbh Procede et entraineurs pour la chromatographie par permeation de gel

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Publication number Priority date Publication date Assignee Title
WO1998040502A1 (fr) 1997-03-14 1998-09-17 Life Technologies, Inc. Transfections activees par des peptides
US6376248B1 (en) 1997-03-14 2002-04-23 Life Technologies, Inc. Peptide-enhanced transfections
US9358300B2 (en) 1998-11-12 2016-06-07 Life Technologies Corporation Transfection reagents
WO2004003542A1 (fr) * 2002-06-28 2004-01-08 Amersham Biosciences Ab Matrices de base a surface modifiee
US7423070B2 (en) 2002-06-28 2008-09-09 Ge Healthcare Bio-Sciences Ab Surface-modified base matrices
US9303098B2 (en) 2008-05-30 2016-04-05 Merck Patent Gmbh Ce(IV)-initiated graft polymerisation on polymers containing no hydroxyl groups
US11369096B2 (en) 2012-04-20 2022-06-28 Commonwealth Scientific And Industrial Research Organisation Process for using crispr to transfect primordial germ cells in avians
US10897881B2 (en) 2012-04-20 2021-01-26 Commonwealth Scientific And Industrial Research Organisation Method of making a chicken with germ cells expressing marker protein
EP3460065A1 (fr) 2012-04-20 2019-03-27 Commonwealth Scientific and Industrial Research Organisation Procédé de transfection cellulaire
TWI609717B (zh) * 2012-11-01 2018-01-01 麥克專利有限公司 多孔性基質擔體之表面改質
CN104736236B (zh) * 2012-11-01 2017-06-13 默克专利股份公司 多孔基体支持物的表面修饰
US10166527B2 (en) 2012-11-01 2019-01-01 Merck Patent Gmbh Surface modification of porous base supports
JP2016502457A (ja) * 2012-11-01 2016-01-28 メルク パテント ゲゼルシャフト ミット ベシュレンクテル ハフツングＭｅｒｃｋ Ｐａｔｅｎｔ Ｇｅｓｅｌｌｓｃｈａｆｔ ｍｉｔ ｂｅｓｃｈｒａｅｎｋｔｅｒ Ｈａｆｔｕｎｇ 多孔質基材支持体の表面修飾
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KR20150082436A (ko) * 2012-11-01 2015-07-15 메르크 파텐트 게엠베하 다공성 기재 지지체의 표면 개질
WO2014067605A1 (fr) * 2012-11-01 2014-05-08 Merck Patent Gmbh Modification superficielle de supports de base poreux
US10195280B2 (en) 2014-07-15 2019-02-05 Life Technologies Corporation Compositions and methods for efficient delivery of molecules to cells
US10792362B2 (en) 2014-07-15 2020-10-06 Life Technologies Corporation Compositions and methods for efficient delivery of molecules to cells
US11872285B2 (en) 2014-07-15 2024-01-16 Life Technologies Corporation Compositions and methods for efficient delivery of molecules to cells
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