CS225197B1 - Preparation of immunosorbents - Google Patents
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- CS225197B1 CS225197B1 CS322082A CS322082A CS225197B1 CS 225197 B1 CS225197 B1 CS 225197B1 CS 322082 A CS322082 A CS 322082A CS 322082 A CS322082 A CS 322082A CS 225197 B1 CS225197 B1 CS 225197B1
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- 239000003547 immunosorbent Substances 0.000 title claims description 11
- 238000002360 preparation method Methods 0.000 title description 6
- 150000004676 glycans Chemical class 0.000 claims description 40
- 229920001282 polysaccharide Polymers 0.000 claims description 39
- 239000005017 polysaccharide Substances 0.000 claims description 39
- 244000005700 microbiome Species 0.000 claims description 12
- 241000222122 Candida albicans Species 0.000 claims description 8
- 241000222178 Candida tropicalis Species 0.000 claims description 7
- 239000002245 particle Substances 0.000 claims description 7
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 5
- 229940095731 candida albicans Drugs 0.000 claims description 5
- 239000003513 alkali Substances 0.000 claims description 4
- 238000004519 manufacturing process Methods 0.000 claims description 3
- 239000006185 dispersion Substances 0.000 claims description 2
- 230000003472 neutralizing effect Effects 0.000 claims description 2
- 229920000057 Mannan Polymers 0.000 description 11
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 11
- 238000000034 method Methods 0.000 description 9
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- 229920000168 Microcrystalline cellulose Polymers 0.000 description 4
- 208000031888 Mycoses Diseases 0.000 description 4
- 238000004132 cross linking Methods 0.000 description 4
- LUEWUZLMQUOBSB-GFVSVBBRSA-N mannan Chemical class O[C@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@@H](O[C@@H]2[C@H](O[C@@H](O[C@H]3[C@H](O[C@@H](O)[C@@H](O)[C@H]3O)CO)[C@@H](O)[C@H]2O)CO)[C@H](O)[C@H]1O LUEWUZLMQUOBSB-GFVSVBBRSA-N 0.000 description 4
- 235000019813 microcrystalline cellulose Nutrition 0.000 description 4
- 239000008108 microcrystalline cellulose Substances 0.000 description 4
- 229940016286 microcrystalline cellulose Drugs 0.000 description 4
- 230000001717 pathogenic effect Effects 0.000 description 4
- 239000002028 Biomass Substances 0.000 description 3
- 241000222120 Candida <Saccharomycetales> Species 0.000 description 3
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 description 3
- 230000007423 decrease Effects 0.000 description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 3
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 2
- BRLQWZUYTZBJKN-UHFFFAOYSA-N Epichlorohydrin Chemical compound ClCC1CO1 BRLQWZUYTZBJKN-UHFFFAOYSA-N 0.000 description 2
- 229920001503 Glucan Polymers 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 238000004873 anchoring Methods 0.000 description 2
- 210000004027 cell Anatomy 0.000 description 2
- 229920002678 cellulose Polymers 0.000 description 2
- 239000001913 cellulose Substances 0.000 description 2
- 235000010980 cellulose Nutrition 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 239000003431 cross linking reagent Substances 0.000 description 2
- 238000002955 isolation Methods 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 238000001556 precipitation Methods 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- 210000002966 serum Anatomy 0.000 description 2
- 239000002594 sorbent Substances 0.000 description 2
- 206010007134 Candida infections Diseases 0.000 description 1
- 241000222173 Candida parapsilosis Species 0.000 description 1
- WQZGKKKJIJFFOK-QTVWNMPRSA-N D-mannopyranose Chemical compound OC[C@H]1OC(O)[C@@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-QTVWNMPRSA-N 0.000 description 1
- 229920000926 Galactomannan Polymers 0.000 description 1
- 229920002581 Glucomannan Polymers 0.000 description 1
- PWHULOQIROXLJO-UHFFFAOYSA-N Manganese Chemical compound [Mn] PWHULOQIROXLJO-UHFFFAOYSA-N 0.000 description 1
- 241000235048 Meyerozyma guilliermondii Species 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 230000009830 antibody antigen interaction Effects 0.000 description 1
- 239000000427 antigen Substances 0.000 description 1
- 230000000890 antigenic effect Effects 0.000 description 1
- 102000036639 antigens Human genes 0.000 description 1
- 108091007433 antigens Proteins 0.000 description 1
- 239000013060 biological fluid Substances 0.000 description 1
- 239000012620 biological material Substances 0.000 description 1
- 229920001222 biopolymer Polymers 0.000 description 1
- 201000003984 candidiasis Diseases 0.000 description 1
- 235000009508 confectionery Nutrition 0.000 description 1
- 230000018044 dehydration Effects 0.000 description 1
- 238000006297 dehydration reaction Methods 0.000 description 1
- 231100000676 disease causative agent Toxicity 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 230000008105 immune reaction Effects 0.000 description 1
- 230000003053 immunization Effects 0.000 description 1
- 238000002649 immunization Methods 0.000 description 1
- 239000012978 lignocellulosic material Substances 0.000 description 1
- 229920002521 macromolecule Polymers 0.000 description 1
- 229910052748 manganese Inorganic materials 0.000 description 1
- 239000011572 manganese Substances 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 230000002906 microbiologic effect Effects 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
- 150000004804 polysaccharides Polymers 0.000 description 1
- 239000011541 reaction mixture Substances 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 230000000405 serological effect Effects 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 230000008961 swelling Effects 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
- 210000005253 yeast cell Anatomy 0.000 description 1
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- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Polysaccharides And Polysaccharide Derivatives (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Description
Vynález sa týká spósobu přípravy zakotvených imunosorbentov z polysacharidov izolovaných z kvasiniek.The invention relates to a process for the preparation of anchored immunosorbents from polysaccharides isolated from yeast.
Hlavnou imunologicky aktívnou zložkou kvasiniek a kvasinkových mikroorganizmov sú manány, alebo manózu obsahujúce heteropolysacharidy, ako galaktomanány, glukomanány, glykurónoxylomanány, ktoré sa nachádzajú na povrchu buniek týchto mikroorganizmov. Manány patogénnych druhov rodu Candida sú vysokovetvené polysacharidy, ktoré majú v hlavnou reťazci a (1 ->· 6), v bočných reťazcoch a (1 -> 2), připadne α (1 3) glykozidické vazby. Imunodominantnou skupinou týchto antigénov sú bočné reťazce skladajúce sa z 5 až 6 manánových jednotiek viazaných a (1 -> 2) glykozidickými vazbami [S. Suzuki a H. Sunayama, Jap. J. Microbiol. 13, 95 (1969)].The major immunologically active component of yeast and yeast microorganisms are mannans, or mannose containing heteropolysaccharides, such as galactomannans, glucomannans, glycuronoxylomannans, which are found on the cell surface of these microorganisms. The mananas of the pathogenic Candida species are high-branched polysaccharides having a (1 → 6) in the backbone, and (1 → 2) in the side chains, or α (1 3) glycosidic bonds. The immunodominant group of these antigens are side chains consisting of 5 to 6 mannan units linked and (1 → 2) glycosidic bonds [S. Suzuki and H. Sunayama, Jap. J. Microbiol. 13, 95 (1969)].
Tieto imunologicky aktivně polysacharidy mikroorganizmov sa móžu izolovat zo živného média niekoíkonásobným zrážaním etanolom, z biomasy kvasiniek extrakciou s hydroxidom draselným o koncentrácii 2 % hmot. [P. A. J. Gorin a J. F. T. Spencer: Adv. Appl. Microbiol. 13, 15, (1970)] alebo s 0,2 mól chloridům sodným [O. Sikl a spol.: Coll. Czechoslov. Chem. Commun. 35, 2969, (1970)] pri zvýšenej teplote. Čisté manány sa získavajú následným niekoíkonásobným zrážaním rozpustných polysacharidov Fehlingovým roztokom. Výtažky imunologicky aktívnych manánov sú však vel'mi nízké a vzhladom na póvodnú biomasu nepresahujú 5 % hmot.These immunologically active polysaccharides of microorganisms can be isolated from the nutrient medium by multiple precipitation with ethanol, from yeast biomass by extraction with potassium hydroxide at a concentration of 2% by weight. [J. A. J. Gorin and J. F. T. Spencer: Adv. Appl. Microbiol. 13, 15, (1970)] or with 0.2 mol of sodium chloride [O. Sikl et al., Coll. Czechloslov. Chem. Commun. 35, 2969, (1970)] at elevated temperature. Pure mannans are obtained by subsequent precipitation of soluble polysaccharides with Fehling's solution several times. However, the yields of immunologically active mannans are very low and do not exceed 5% by weight relative to the original biomass.
Kedže manány kvasinkových mikroorganizmov sú vo vodě dobré rozpustné, nedajú sa preto priamo použit na přípravu pevných sorbentov. Aby sa získali vhodné nerozpustné imunosorbenty, je potřebné polysacharidy z kvasinkovitých mikroorganizmov vhodné modifikovat sieťovaním [E. Kuniak, R. H. Marchessault, Starch 24, 110, (1972)], a to tak, aby imunodominantné skupiny polysacharidu ostali po sletovaní zachované pre imunosorbciu špecifických protilátok z krvného séra. Molekulová hmotnost manánov polysacharidov kvasinkovitých mikroorganizmov je relativné nízká a pohybuje sa medzi 15 000 až 30 000 daltonov. Na jednej molekule polysacharidu sa nachádza niekol'ko determlnantných skupin, avšak len časť z nich sa zúčastňuje špecifickej imunologické j reakcie antigén — protilátka. (J. Šandula a M. Vojková, Folia Microhiologica, 19, 94, 1974). Zosieťovaním manánu dochádza ešte k hustejšiemu rozloženiu determinantných skupin v makromolekule manánu, takže časť aktvínych centier imunosorbenta sa nezúčastňuje interakcie antigén — protilátka z priestorových dóvodov.Since the manganese of yeast microorganisms are well soluble in water, they cannot be used directly for the preparation of solid sorbents. In order to obtain suitable insoluble immunosorbents, polysaccharides from yeast microorganisms need to be modified by cross-linking [E. Kuniak, R. H. Marchessault, Starch 24, 110, (1972)], so that the immunodominant polysaccharide groups remain after immunization to immunosorb the specific antibodies from the blood serum. The molecular weight of the mannan polysaccharides of yeast microorganisms is relatively low and is between 15,000 and 30,000 daltons. Several determinant groups are present on a single polysaccharide molecule, but only a fraction of them participate in a specific antigen-antibody immunological reaction. (J. Sandula and M. Vojkova, Folia Microhiologica, 19, 94, 1974). Crosslinking of mannan results in an even denser distribution of the determinant groups in the mannan macromolecule, so that part of the active immunosorbent centers do not participate in the antigen-antibody interaction for spatial reasons.
Imunologicky aktivně manány sa vyskytujú v buňkových stěnách kvasiniek v komplexe s bielkovinami a imunologicky neaktívnym glukanom (C. E. Ballau, Adv. Microbiol. Phys. 14, 93, 1976). Úplné odstránenie týchto sprievodných biopolymérov naráža na tažkosti a vyžaduje zložitý purifikačný proces. Zistili sme, že pre přípravu účinného imunosorbentu nie je potřebné manán úplné purifikovať, malé množstvo bielkoviny a glukánu nie je na závadu. Purififikačnými stupňami jednak klesá výtažok polysacharidu, klesá jeho molekulová hmotnost, čo sťažuje účinné zosieťovanie polysacharidu a nie zanedbatelnou mierou klesá počet imunodominantných skupin schopných viazať homológne protilátky, preto je výhodnejšie použiť polysacharid len čiastočne puntíkovaný s vyššou molekulovou hmotnosťou.Immunologically active mannans occur in yeast cell walls complexed with proteins and immunologically inactive glucan (C.E. Ballau, Adv. Microbiol. Phys. 14, 93, 1976). The complete removal of these accompanying biopolymers encounters difficulties and requires a complex purification process. We have found that for the preparation of an effective immunosorbent, it is not necessary to purify mannan completely, a small amount of protein and glucan is not a problem. On the one hand, the purification steps decrease the polysaccharide yield, decrease its molecular weight, which makes it difficult to cross-link the polysaccharide efficiently and not to a negligible extent the number of immunodominant groups capable of binding homologous antibodies decreases, therefore it is preferable to use only partially purified polysaccharide.
Podstata vynálezu spočívá v tom, že polysacharid izolovaný z kvasinkovitých mikroorganizmov, s výhodou Candida albicans alebo Candida tropicalis sa v prvom stupni zosieťuje s l-chlór-2,3-epoxipropanom v množstve 15 až 30 % hmot. na hmotnost polysacharidu v alkalickom prostředí a v druhom stupni sa zosieťovaný polysacharidový gél podrobí vodnej dispergácii vo vysokoobrátkovom mixéri za súčasnej neutralizácie alkálií a v treťom stupni sa premyté nerozpustné polysacharidové částice odvodnia na filtri alkoholom. a vysušia pri teplote do 60° Celsia.According to the invention, the polysaccharide isolated from yeast microorganisms, preferably Candida albicans or Candida tropicalis, is crosslinked in the first step with 1-chloro-2,3-epoxipropane in an amount of 15 to 30% by weight. to the weight of the polysaccharide in an alkaline medium and in a second stage, the cross-linked polysaccharide gel is subjected to an aqueous dispersion in a high-shear mixer while neutralizing the alkali and in a third stage the washed insoluble polysaccharide particles are dewatered on the filter with alcohol. and dried at a temperature of up to 60 ° C.
Takto připravený nerozpustný polysacharid s velkosťou častíc 80 až 250 mikrónov je vhodným imunosorbentom pre izoláciu protilátok proti róznym hubovitým ochoreniam.The thus prepared insoluble polysaccharide having a particle size of 80 to 250 microns is a suitable immunosorbent for isolation of antibodies against various fungal diseases.
Najčastejším póvodcom mykotických ochorení u l'udí je Candida albicans, na ktoré připadá výše 80 % patologických mikrobiologických nálezov. Zbytok tvoria mikroorganizmy ako Candida tropicalis, C. stellatoidea, C. guilliermOndii, C. parapsilosis, C. fanata a iné. Mnohé druhy Candida majú totožné alebo podobné antigénne zloženie ako C. albicans, sérologicky sú ťažko rozlišitelné a na mikroorganizmy tejto sérologickej skupiny připadá až 90 % všetkých kandidóz. Vzhladom na to, že mnohé kmene Candida albicans sú silno patogénne, tak ich kultivácia za účelom přípravy imunologicky aktívnych polysacharidov sa musí robiť za přísné sterilných podmienok, čo zvyšuje náklady na přípravy biomasy.The most common causative agent of fungal diseases in humans is Candida albicans, which accounts for over 80% of pathological microbiological findings. The rest consists of microorganisms such as Candida tropicalis, C. stellatoidea, C. guilliermOndii, C. parapsilosis, C. fanata and others. Many Candida species have the same or similar antigenic composition as C. albicans, are serologically difficult to distinguish, and microorganisms of this serological group account for up to 90% of all candidiasis. Because many strains of Candida albicans are strongly pathogenic, their cultivation to produce immunologically active polysaccharides must be done under strict sterile conditions, increasing the cost of biomass production.
Je preto výhodné pre tieto účely použiť kmen Candida tropicalis, ktorého sa priemyslovo využívá na výrobu krmného droždia, je patogénny alebo len málo patogénny a naviac dobré sa kultivuje na roznych odpadných lignocelulózových materiáloch. Manán z C. tropicalis použitý ako imunosorbent zachytí protilátky séra homológneho druhu ako aj špecifické látky vytvořené mikroorganizmami voči manánom uvedenej séroskupiny.It is therefore advantageous to use for this purpose the Candida tropicalis strain, which is industrially used for the production of feed yeast, is pathogenic or only slightly pathogenic and, moreover, it is cultivated well on various waste lignocellulosic materials. C. tropicalis mannan used as an immunosorbent will capture serum antibodies of a homologous species as well as specific substances produced by microorganisms against the mannan of said serogroup.
Zakotvenie imunoaktívneho polysacharidu je výhodné prevádzať tak, že imunoaktívny polysacharid sa rozpúšťa v hydroxide sodnom o koncentrácii 5 až 8 % hmot., tak, aby koncentrácia polysacharidu bola 15 až 30 percent hmot. a takýto roztok sa zmieša s potřebným množstvom mikrokryštalickej celulózy s velkosťou častíc 50 až 250 μπι. Roztok sa nechá asi Ví hod. dobré nasiaknuť do celulózy a až potom sa přidá sieťovací reagent. Sieťovanie sa prevádza najprv pri teplote 20 až 30 °C, potom pri 50 °C, pri ktorej teplote sa reakcia ukončí. Před defibráciou zosieteného gélu je výhodné pridať kyselinu octovú na neutralizáciu volných alkálií. Bez neutralizácie by sa získali gólové částice velmi malých rozmerov, ktoré by neboli vhodné pre prácu v koloně.The anchoring of the immunoactive polysaccharide is preferably carried out by dissolving the immunoactive polysaccharide in sodium hydroxide at a concentration of 5-8% by weight, such that the polysaccharide concentration is 15-30% by weight. and such a solution is mixed with the necessary amount of microcrystalline cellulose having a particle size of 50 to 250 μπι. The solution is left for about 1 hour. soak up the cellulose before adding the crosslinking reagent. The crosslinking is carried out first at 20 to 30 ° C, then at 50 ° C at which temperature the reaction is complete. Prior to defibrating the crosslinked gel, it is preferred to add acetic acid to neutralize the free alkali. Without neutralization, goal particles of very small dimensions would be obtained which would not be suitable for column work.
Hlavnou výhodou nového postupu je skutočnosť, že sa získá imunoaktívny sorbent s 2- až 3násobne vyšším Specifickým povrchom, t. j. zvýši sa počet aktívnych imunodominantných skupin schopných reakcie antigén-protilátka. Táto skutočnosť je významná vzhladom na to, že izolácia i nepurifikovaného imunoaktívneho polysacharidu a nákladnejšia ako zakotvenie polysacharidu na relativné lacnom nosiči. Ďalšou výhodou tohto postupu je to, že zakotvený imunoaktívny gél na celulózovom nosiči má lepšie reologické vlastnosti pri práci v koloně.The main advantage of the new process is that an immunoactive sorbent is obtained with a 2- to 3-fold higher specific surface area, i. j. the number of active immunodominant groups capable of antigen-antibody response will increase. This is significant because the isolation of even the unpurified immunoactive polysaccharide is more expensive than anchoring the polysaccharide on a relatively inexpensive carrier. Another advantage of this procedure is that the anchored immunoactive gel on the cellulose carrier has better rheological properties when working in the column.
Ďalšie přednosti a detaily nového postupu sú zřejmé z príkladovej časti postupu, ktorá nie je vyčerpávajúca, ale len informatívna na demonštráciu podstatných znakov nového postupu přípravy zakotveného imunosorbenta vhodného na izolovanie protilátok proti mykotickým ochoreniam spósobeným kvasinkovitými mikroorganizmami rodu Candida.Further advantages and details of the novel process are evident from the non-exhaustive, but non-exhaustive, part of the process to demonstrate the essential features of the novel process for the preparation of an anchored immunosorbent suitable for isolating antibodies against fungal diseases caused by yeast Candida microorganisms.
Přikladl g surového, suchého polysacharidu izolovaného z buniek kvasinkovitých mikroorganizmov Candida tropicalis sa zvlhčí s 20 ml destilovanej vody a nechá sa 5 až 10 min. napučiavať. Potom sa přidá 4 ml hydroxidu sodného o koncentrácii 40 % hmot., dobré sa zamieša a nechá rozpúšťať, až je roztok homogénny bez zjavných nerozpustných častíc, čo obyčajne netrvá viac ako 10 až 15 min. Do alkalického roztoku polysacharidu sa naraz přidá 10 g mikrokryštalickej celulózy o velkosti častíc 50 až 250 ,um a dobré sa zamieša tak, aby roztok polysacharidu sa rovnoměrně nasal do mikrokryštalickej celulózy, čo tvoří asi 20 až 30 min. Potom sa přidá 2 ml l-chlór-2,3-epoxipropánu (epichlórhydrín) a dobré sa zamieša tak, aby sieťovací reagent bol homogenně distribuovaný v celom objeme reakčnej zmesi. Reakčnú banku uzatvoríme a necháme reagovat pri laboratórnej teplote 1 až 2 hod., potom zvýšíme teplotu postupné až na 50 °C a pri tejto teplote necháme reakčnú zmes reagovat 1 hod., načo zosietený polysacharidový gél mixujeme vo voděExample 1 g of crude, dry polysaccharide isolated from cells of yeast Candida tropicalis is moistened with 20 ml of distilled water and left for 5-10 min. swell. Then 4 ml of 40% sodium hydroxide is added, mixed well and allowed to dissolve until the solution is homogeneous without apparent insoluble particles, which usually does not take more than 10 to 15 minutes. 10 g of microcrystalline cellulose having a particle size of 50 to 250 µm is added to the alkaline polysaccharide solution at once and mixed well so that the polysaccharide solution is uniformly aspirated into the microcrystalline cellulose, which is about 20 to 30 minutes. Then 2 ml of 1-chloro-2,3-epoxipropane (epichlorohydrin) is added and mixed well so that the crosslinking agent is homogeneously distributed throughout the volume of the reaction mixture. Seal the reaction flask and react at room temperature for 1 to 2 hours, then raise the temperature gradually up to 50 ° C and react at this temperature for 1 hour, then mix the cross-linked polysaccharide gel in water
225 (do ktorej sa přidá kyselina octová na neutralizační volných alkáliíj asi 30 až 60 s v rýchloobrátkovom mixéri. Nerozpustný zakotvený polysacharidový gél sa niekolkonásobne dekantuje od najjemnejších podielov, ktoré by velmi zvyšovali odpor pri práci na kolóne. Dekantované a dokonalé premyté gélové částice sa na filtri postupné odvodnia alkoholom, resp. acetónom a vysušia pri teplote do 60 °C. Napúčací objem připraveného zakotveného imunosorbenta je okolo 6 až 7 ml/g a je vhodný pre imunosorbciu protilátok z biologických tekutin.225 (to which acetic acid is added to neutralize the free alkali is about 30 to 60 s in a quick mixer. The insoluble ground polysaccharide gel is decanted several times from the finest fractions that would greatly increase the resistance when working on the column. Gradual dehydration with alcohol or acetone and drying at temperatures up to 60 ° C. The swelling volume of the prepared grounded immunosorbent is about 6-7 ml / g and is suitable for immunosorbing antibodies from biological fluids.
Příklad 2Example 2
Postup podl'a příkladu 1 s tým rozdielom, že na sieťovanie použijeme 4 ml epichlorhydrínu a rozpustný polysacharid zakotvíme na 30 g mikrokryštalickej celulózy. Vlastnosti získaného zakotveného imunolo9 7 gicky aktívneho polysacharidového gélu sú podobné ako u příkladu 1.The procedure of Example 1 was followed except that 4 ml of epichlorohydrin was used for crosslinking and the soluble polysaccharide was anchored to 30 g of microcrystalline cellulose. The properties of the obtained grounded immunologically active polysaccharide gel are similar to those of Example 1.
Příklad 3Example 3
Postup podl'a příkladu 1 s tým rozdielom, že ako imunologicky aktívny polysacharid sa použije polysacharid izolovaný z mikroorganizmu Candida albicans. Zosietený, zakotvený polysacharid má obdobné vlastnosti ako zosietený polysacharid izolovaný z Candida tropicalis.The process of Example 1 except that the immunologically active polysaccharide used is a polysaccharide isolated from Candida albicans. The cross-linked, grounded polysaccharide has similar properties to the cross-linked polysaccharide isolated from Candida tropicalis.
Popísaná metoda přípravy imunosorbentov z mikrobiálnych polysacharidov je univerzálna, může byť použitá nielen pre kandidy, ale pre lubovolné imunologicky aktivně polysacharidy.The described method of preparation of immunosorbents from microbial polysaccharides is universal, it can be used not only for candies but for any immunologically active polysaccharides.
Vynález má použitie v zdravotníctve pri dokáže, izolácii a purifikácii účinných protilátok z biologického materiálu pri mykotických ochoreniach ako i v alergológii.The invention is of use in the medical field to detect, isolate and purify effective antibodies from biological material in fungal diseases as well as in allergology.
Claims (1)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CS322082A CS225197B1 (en) | 1982-05-05 | 1982-05-05 | Preparation of immunosorbents |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CS322082A CS225197B1 (en) | 1982-05-05 | 1982-05-05 | Preparation of immunosorbents |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CS225197B1 true CS225197B1 (en) | 1984-02-13 |
Family
ID=5371655
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CS322082A CS225197B1 (en) | 1982-05-05 | 1982-05-05 | Preparation of immunosorbents |
Country Status (1)
| Country | Link |
|---|---|
| CS (1) | CS225197B1 (en) |
-
1982
- 1982-05-05 CS CS322082A patent/CS225197B1/en unknown
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