JPH07285885A - Method for producing immunoglobulin - Google Patents
Method for producing immunoglobulinInfo
- Publication number
- JPH07285885A JPH07285885A JP10499194A JP10499194A JPH07285885A JP H07285885 A JPH07285885 A JP H07285885A JP 10499194 A JP10499194 A JP 10499194A JP 10499194 A JP10499194 A JP 10499194A JP H07285885 A JPH07285885 A JP H07285885A
- Authority
- JP
- Japan
- Prior art keywords
- immunoglobulin
- whey
- exchange resin
- cation exchange
- concentration
- Prior art date
- Legal status (The legal status 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 status listed.)
- Pending
Links
- 108060003951 Immunoglobulin Proteins 0.000 title claims abstract description 32
- 102000018358 immunoglobulin Human genes 0.000 title claims abstract description 32
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 10
- 239000005862 Whey Substances 0.000 claims abstract description 18
- 102000007544 Whey Proteins Human genes 0.000 claims abstract description 18
- 108010046377 Whey Proteins Proteins 0.000 claims abstract description 18
- 238000000108 ultra-filtration Methods 0.000 claims abstract description 17
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 claims abstract description 14
- 239000000178 monomer Substances 0.000 claims abstract description 9
- NWUYHJFMYQTDRP-UHFFFAOYSA-N 1,2-bis(ethenyl)benzene;1-ethenyl-2-ethylbenzene;styrene Chemical compound C=CC1=CC=CC=C1.CCC1=CC=CC=C1C=C.C=CC1=CC=CC=C1C=C NWUYHJFMYQTDRP-UHFFFAOYSA-N 0.000 claims abstract description 8
- 239000003729 cation exchange resin Substances 0.000 claims abstract description 8
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 claims abstract description 7
- 239000004471 Glycine Substances 0.000 claims abstract description 7
- 238000000926 separation method Methods 0.000 claims abstract description 6
- 239000005018 casein Substances 0.000 claims abstract description 3
- BECPQYXYKAMYBN-UHFFFAOYSA-N casein, tech. Chemical compound NCCCCC(C(O)=O)N=C(O)C(CC(O)=O)N=C(O)C(CCC(O)=N)N=C(O)C(CC(C)C)N=C(O)C(CCC(O)=O)N=C(O)C(CC(O)=O)N=C(O)C(CCC(O)=O)N=C(O)C(C(C)O)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=O)N=C(O)C(CCC(O)=O)N=C(O)C(COP(O)(O)=O)N=C(O)C(CCC(O)=N)N=C(O)C(N)CC1=CC=CC=C1 BECPQYXYKAMYBN-UHFFFAOYSA-N 0.000 claims abstract description 3
- 235000021240 caseins Nutrition 0.000 claims abstract description 3
- 230000002255 enzymatic effect Effects 0.000 claims abstract description 3
- 238000001556 precipitation Methods 0.000 claims abstract description 3
- 239000001509 sodium citrate Substances 0.000 claims abstract description 3
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 claims abstract description 3
- 229920000642 polymer Polymers 0.000 claims description 6
- 102000006395 Globulins Human genes 0.000 claims 1
- 108010044091 Globulins Proteins 0.000 claims 1
- 239000000427 antigen Substances 0.000 abstract description 3
- 102000036639 antigens Human genes 0.000 abstract description 3
- 108091007433 antigens Proteins 0.000 abstract description 3
- 241000219977 Vigna Species 0.000 abstract 1
- 235000010726 Vigna sinensis Nutrition 0.000 abstract 1
- 238000005469 granulation Methods 0.000 abstract 1
- 230000003179 granulation Effects 0.000 abstract 1
- 229940072221 immunoglobulins Drugs 0.000 abstract 1
- 238000000034 method Methods 0.000 description 15
- 238000002360 preparation method Methods 0.000 description 7
- 241000283690 Bos taurus Species 0.000 description 4
- 210000003022 colostrum Anatomy 0.000 description 4
- 235000021277 colostrum Nutrition 0.000 description 4
- 235000018102 proteins Nutrition 0.000 description 4
- 102000004169 proteins and genes Human genes 0.000 description 4
- 108090000623 proteins and genes Proteins 0.000 description 4
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 3
- 235000013305 food Nutrition 0.000 description 3
- 235000013336 milk Nutrition 0.000 description 3
- 239000008267 milk Substances 0.000 description 3
- 210000004080 milk Anatomy 0.000 description 3
- 239000012465 retentate Substances 0.000 description 3
- 102000008192 Lactoglobulins Human genes 0.000 description 2
- 108010060630 Lactoglobulins Proteins 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 238000001042 affinity chromatography Methods 0.000 description 2
- 238000005194 fractionation Methods 0.000 description 2
- 238000002955 isolation Methods 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 101001008231 Bos taurus Beta-lactoglobulin Proteins 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- 101000798114 Homo sapiens Lactotransferrin Proteins 0.000 description 1
- 102000004407 Lactalbumin Human genes 0.000 description 1
- 108090000942 Lactalbumin Proteins 0.000 description 1
- 102000010445 Lactoferrin Human genes 0.000 description 1
- 108010063045 Lactoferrin Proteins 0.000 description 1
- 229940098773 bovine serum albumin Drugs 0.000 description 1
- 239000000872 buffer Substances 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 239000007979 citrate buffer Substances 0.000 description 1
- 239000012141 concentrate Substances 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 239000006071 cream Substances 0.000 description 1
- 235000013365 dairy product Nutrition 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000002523 gelfiltration Methods 0.000 description 1
- 102000050459 human LTF Human genes 0.000 description 1
- 235000021244 human milk protein Nutrition 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 238000005342 ion exchange Methods 0.000 description 1
- CSSYQJWUGATIHM-IKGCZBKSSA-N l-phenylalanyl-l-lysyl-l-cysteinyl-l-arginyl-l-arginyl-l-tryptophyl-l-glutaminyl-l-tryptophyl-l-arginyl-l-methionyl-l-lysyl-l-lysyl-l-leucylglycyl-l-alanyl-l-prolyl-l-seryl-l-isoleucyl-l-threonyl-l-cysteinyl-l-valyl-l-arginyl-l-arginyl-l-alanyl-l-phenylal Chemical group C([C@H](N)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CS)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)NCC(=O)N[C@@H](C)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CS)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(O)=O)C1=CC=CC=C1 CSSYQJWUGATIHM-IKGCZBKSSA-N 0.000 description 1
- 235000021242 lactoferrin Nutrition 0.000 description 1
- 229940078795 lactoferrin Drugs 0.000 description 1
- 239000003446 ligand Substances 0.000 description 1
- 150000002632 lipids Chemical group 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 238000005185 salting out Methods 0.000 description 1
- 230000001235 sensitizing effect Effects 0.000 description 1
- 238000010008 shearing Methods 0.000 description 1
- 235000020183 skimmed milk Nutrition 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
- 235000021241 α-lactalbumin Nutrition 0.000 description 1
Landscapes
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
- Separation Using Semi-Permeable Membranes (AREA)
- Peptides Or Proteins (AREA)
Abstract
(57)【要約】
【目的】 免疫グロブリン及びこれによつて得られた免
疫グロブリン（抗原）による免疫グロブリン単量体の製
造法に関する。
【構成】 カゼインの等電点（ｐＨ４．５−４．６）沈
殿又はレンニン等の酵素処理で得た乳清に、エチレンジ
アミン四酢酸の濃度０．５−１０ｍＭ、又はグリシンの
濃度が５０−５００ｍＭになるように加え、クエン酸ナ
トリウム溶液で乳清のｐＨを６．０−６．５に調整し、
これに陽イオン交換樹脂を接触させ、５０，０００又は
１００，０００ダルトンの分離限界を有する限外濾過モ
ジユールを用いて、目的とする免疫グロブリンを得る。(57) Abstract: OBJECTIVE manufacturing <br/> granulation for immunoglobulin monomer by immunoglobulins and resultant immunoglobulin Te cowpea to (antigen) relates. [Constitution] The whey obtained by the isoelectric point (pH 4.5-4.6) precipitation of casein or the enzymatic treatment of rennin, etc., was added with ethylenediaminetetraacetic acid at a concentration of 0.5-10 mM or glycine at a concentration of 50-500 mM. And adjust the pH of the whey to 6.0-6.5 with a sodium citrate solution,
This is contacted with a cation exchange resin, and an immunoglobulin of interest is obtained by using an ultrafiltration module having a separation limit of 50,000 or 100,000 daltons.
Description
【０００１】[0001]
【産業上の利用分野】この発明は免疫グロブリン及びこ
れによつて得られた免疫グロブリンによる免疫グロブリ
ン単量体の製造法に関する。BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an immunoglobulin and a method for producing an immunoglobulin monomer using the immunoglobulin thus obtained.
【０００２】[0002]
【従来の技術】ウシ初乳には１ml当たりIgGl 64.9、IgG
2 2.2、IgA 3.5、IgM 8.7mgの免疫グロブリン(Ig)が含
まれており(Butler, J.E., ed. Butler, J.E., “The R
uminantImmune System", pp3-55, Plenum Press, New Y
ork, 1981)、このIgを分取し、積極的に用いようとの試
みが多方面で検討されている。2. Description of the Related Art Bovine colostrum contains IgGl 64.9, IgG1 per ml.
2 2.2, IgA 3.5, IgM 8.7 mg of immunoglobulin (Ig) is included (Butler, JE, ed. Butler, JE, “The R
uminantImmune System ", pp3-55, Plenum Press, New Y
ork, 1981), attempts to sort out this Ig and use it positively have been studied in various fields.
【０００３】Igの中でもメインタ−ゲツトにされている
のは、最も量の多いIgG画分であり、これの一般的な調
製法は、塩析、ゲル濾過、イオン交換法等の組み合わせ
であり、特異性が要求される場合にはプロテインＡをリ
ガンドにしたアフイニテイ−クロマトグラフイ−を用い
ている。特殊な抗原に対する抗体を得、研究又は医薬品
とするのであれば、これらの調製法によるコストの上昇
を吸収し得るが、食品又はそれに類する物に用いるので
あれば、それら調製法による製品のコストは非現実的で
ある。Of the Igs, the main target is the IgG fraction with the highest amount, and the general preparation method for this is a combination of salting out, gel filtration, ion exchange, etc. When specificity is required, affinity chromatography using protein A as a ligand is used. If the antibody against a specific antigen is obtained and used for research or medicine, the increase in cost due to these preparation methods can be absorbed, but if it is used for foods or the like, the cost of the product due to these preparation methods is Unrealistic.
【０００４】したがつて、既述の調製法以外にもFeCl
3(Kuwata, T. et. al., Eliminationof β-lactoglobul
in from whey to stimulate human milk protein. j. F
oodSci., 50, 605-609, 1985)、NaCl（Mailliart, P. e
t. al., J. Food Sci., 53(3), 743, 1988)、限外濾過
（桐原 修、ウシ免疫グロブリンの分離と利用、酪農科
学、食品の研究、39、A-301-A-305、1990)等による調製
法も報告されている。Therefore, in addition to the above-mentioned preparation method, FeCl
3 (Kuwata, T. et.al., Eliminationof β-lactoglobul
in from whey to stimulate human milk protein.j.F
oodSci., 50, 605-609, 1985), NaCl (Mailliart, P. e.
t. al., J. Food Sci., 53 (3), 743, 1988), ultrafiltration (Osamu Kirihara, isolation and utilization of bovine immunoglobulin, dairy science, food research, 39, A-301-A) -305, 1990) etc. have been reported.
【０００５】これらの各方法は非常に簡便であり、製造
コストを低く抑えられる利点を有するものの、特異性に
欠け、特に限外濾過(UF)法以外の方法は抗体価の著しい
低下を来す。またUF法は基本的に分子量差による分画で
あり有効な方法であるが、牛乳ホエ−にはβ-ラクトグ
ロブリン、α-ラクトアルブミン、ウシ血清アルブミン
等に代表されるように種々の物質と結合しやすいタンパ
ク質が存在する。Although each of these methods is very simple and has the advantage that the production cost can be kept low, it lacks in specificity, and methods other than the ultrafiltration (UF) method in particular cause a marked decrease in antibody titer. . Further, the UF method is basically an effective method because it is a fractionation based on a difference in molecular weight, but milk whey contains various substances such as β-lactoglobulin, α-lactalbumin, and bovine serum albumin. There are proteins that are easy to bind.
【０００６】とくに既述タンパク質は脂質類と非常に結
合しやすく、またβ-ラクトグロブリンはホエ-中に存在
するラクトフエリンとも強く結合する（Ena, J.M. et.
al.,Isolation of human lactoferrin by affinity chr
omatography using insolubilized bovine β-lactoglo
bulin. J. Chromato., 525, 442-446, 1990)ことから、
ホエ-中でのこれらのタンパク質の見かけ上の分子量は
非常に大きく、UF処理による分画の障害となつている。[0006] In particular, the above-mentioned protein is very easily bound to lipids, and β-lactoglobulin is also strongly bound to lactoferrin existing in whey (Ena, JM et.
al., Isolation of human lactoferrin by affinity chr
omatography using insolubilized bovine β-lactoglo
bulin. J. Chromato., 525, 442-446, 1990)
The apparent molecular weight of these proteins in whey is very high, which impedes the fractionation by UF treatment.
【０００７】したがつて、UF法によりIgを製造した場
合、その純度は70-80%程度(南 善行らの「牛乳中の免疫
グロブリンの濃縮方法」（特開昭60-75433号公報)、ノ
ベルトコ-テらの「乳および/または初乳免疫グロブリン
溶液の製造方法」（特開昭61-68429号公報)であり、種
々の夾雑物を含み純度的に満足できるものではなかつ
た。また従来の方法ではIgGが重合体として存在してい
る場合が多く、その力価を安定にすることは殆んど不可
能に近い状態でもあつた。Accordingly, when Ig is produced by the UF method, its purity is about 70-80% ("A method for concentrating immunoglobulin in milk" by Zen Minamoto et al. (JP-A-60-75433), It is "Production method of milk and / or colostrum immunoglobulin solution" by Novelto Cote et al. (Japanese Patent Laid-Open No. 61-68429), and it is not satisfactory in purity because it contains various impurities. In most cases, IgG was present as a polymer in the method described above, and it was almost impossible to stabilize its titer.
【０００８】[0008]
【発明が解決しようとする課題】この発明は従来の技術
における種々の障害及び問題点を改善しようとするもの
である。SUMMARY OF THE INVENTION The present invention seeks to overcome various obstacles and problems associated with the prior art.
【０００９】[0009]
【課題を解決するための手段】ここにおいてこの発明
は、カゼインの等電点(pH4.5-4.6)沈殿又はレンニン等
の酵素処理で得た乳清に、エチレンジアミン四酢酸の濃
度0.5-10mM、又はグリシンの濃度が50-500mMになるよう
に加え、クエン酸ナトリウム溶液で乳清のpHを6.0-6.5
に調整し、これに陽イオン交換樹脂を接触させ、50,000
又は100,000ダルトンの分離限界を有する限外濾過モジ
ユ−ルを用いて、目的とする免疫グロブリンを得ること
を特徴とする免疫グロブリンの製造法を提案し、更にこ
れにもとづいて、上記方法で得た乳清を100,000ダルト
ンの分離限界を有する限外濾過モジユ−ルで得た免疫グ
ロブリン画分に陽イオン交換樹脂を接触させ、免疫グロ
ブリンの重合体を除去することを特徴とする免疫グロブ
リン単量体の製造法並びに上記方法で得た免疫グロブリ
ンと、陽イオン交換樹脂とを接触させ、免疫グロブリン
重合体を除去することを特徴とする免疫グロブリン単量
体の製造法を提案するものである。Means for Solving the Problems Here, the present invention is the isoelectric point of casein (pH 4.5-4.6) precipitation or whey obtained by enzymatic treatment such as rennin, the concentration of ethylenediaminetetraacetic acid 0.5-10 mM, Alternatively, add glycine to a concentration of 50-500 mM and adjust the whey pH to 6.0-6.5 with sodium citrate solution.
Adjusted to 50,000 and contacted with a cation exchange resin.
Or, using an ultrafiltration module having a separation limit of 100,000 daltons, we have proposed a method for producing an immunoglobulin characterized by obtaining the immunoglobulin of interest, and further based on this, obtained by the above method An immunoglobulin monomer characterized by contacting a cation exchange resin with an immunoglobulin fraction obtained by ultrafiltration module having a separation limit of 100,000 daltons for whey to remove an immunoglobulin polymer. And a method for producing an immunoglobulin monomer, which comprises contacting the immunoglobulin obtained by the above method with a cation exchange resin to remove an immunoglobulin polymer.
【００１０】[0010]
【作用】上記方法によつて免疫グロブリンを得、これに
もとづいて免疫グロブリン単量体を製造する。The immunoglobulin is obtained by the above method, and the immunoglobulin monomer is produced based on the obtained immunoglobulin.
【００１１】[0011]
【実施例】 (1)ウシ初乳乳清の調製:分娩3週間前からヒトIgEを常法
で感作したウシから、分娩後0-3日目までの初乳を採取
した。クリ-ムセパレ-タで脂肪を除去し、脱脂乳を得
た。これに等量の脱イオン水を加え、EDTAを0.5-10mM
(好ましくは1mM)又はグリシン濃度が50-500mM(好ましく
は100mM)になるように加えた後、クエン酸でpHは4.5-4.
6に調整した。生じた沈殿物を遠心分離で除去し、乳清
を得た。この乳清のpHを水酸化ナトリウムで6.0-6.5に
調整し、実験に用いた。この時、感作する抗原(ヒトIg
E)はこの発明を説明するためだけのものである。Examples (1) Preparation of bovine colostrum whey: Colostrum from 0 to 3 days after calving was collected from cows sensitized with human IgE by a conventional method from 3 weeks before calving. The fat was removed with a cream separator to obtain skim milk. Add an equal volume of deionized water to this and add EDTA to 0.5-10 mM.
(Preferably 1 mM) or after adding so that the glycine concentration is 50-500 mM (preferably 100 mM), the pH with citric acid is 4.5-4.
Adjusted to 6. The resulting precipitate was removed by centrifugation to obtain whey. The pH of this whey was adjusted to 6.0-6.5 with sodium hydroxide and used in the experiment. At this time, the sensitizing antigen (human Ig
E) is only for explaining the present invention.
【００１２】(2)IgG単量体の調製:上記(1)で得た乳清30
0mlに、DEAE-セルロフアイン（生化学工業社製）20gを
加え、穏やかに攪拌した後、ブフナ-漏斗で濾過し、非
吸着画分を集め、50K又は100Kダルトンの分離限界を有
する限外濾過モジユ-ル(旭化成社製)を用いて限外濾過
を行なつた。濃縮されたリテンテイトを1mMEDTA又は100
mMグリシンを含む10mMクエン酸緩衝液(pH6.0-6.5)で希
釈し、限外濾過を繰り返した。この操作により低分子物
質を除去した後、リテンテイトを凍結乾燥した。(2) Preparation of IgG monomer: whey 30 obtained in (1) above
To 0 ml, 20 g of DEAE-cellulophane (manufactured by Seikagaku Corporation) was added, gently stirred, and then filtered with a Buchna-funnel to collect non-adsorbed fractions, and an ultrafiltration module having a separation limit of 50 K or 100 K daltons. -Ul (manufactured by Asahi Kasei Corp.) was used for ultrafiltration. Concentrate retentate with 1 mM EDTA or 100
It was diluted with 10 mM citrate buffer (pH 6.0-6.5) containing mM glycine, and ultrafiltration was repeated. After removing low molecular weight substances by this operation, the retentate was freeze-dried.
【００１３】(3)上記(1)で得た乳清を(2)で用いた限外
濾過モジユ-ル及び1mMEDTA又は100mMグリシンを含む緩
衝液存在下で濾過し、得られたリテンテイトとDEAE-セ
ルロフアインを接触させ、非吸着画分を集め、これを凍
結乾燥した。(3) The whey obtained in (1) above is filtered in the presence of the ultrafiltration module used in (2) and a buffer containing 1 mM EDTA or 100 mM glycine to obtain the retentate and DEAE- Cellulophane was contacted, non-adsorbed fractions were collected and freeze-dried.
【００１４】(4)上記(2)に記載の手法でIgG単量体を製
造する場合、限外濾過中のポンプに起因する加圧や剪断
力等の物理的力により、IgGが重合体を形成するのが散
見される。このIgG重合体を(3)に記載の手法のように、
DEAE-セルロフアインを接触させることにより除去し、
凍結乾燥した。(4) When the IgG monomer is produced by the method described in (2) above, the IgG is polymerized by the physical force such as pressurization and shearing force caused by the pump during the ultrafiltration. It is sporadically formed. This IgG polymer, as in the method described in (3),
Removed by contacting DEAE-celluloin,
Lyophilized.
【００１５】次に(2),(3)及び(4)に記載の方法により得
たIgG調整物の収率、純度及び力価をまとめて表１に示
す。Next, the yield, purity and titer of the IgG preparations obtained by the methods described in (2), (3) and (4) are summarized in Table 1.
【００１６】[0016]
【表１】 [Table 1]
【００１７】[0017]
【発明の効果】上記方法によれば、乳清にエチレンジア
ミン四酢酸及びグリシンを加え、pH6.0-6.5において、
陽イオン交換樹脂及び限外濾過モジユ-ルを用いて免疫
グロブリンを得ることができるものである。According to the above method, whey is added with ethylenediaminetetraacetic acid and glycine, at pH 6.0-6.5,
An immunoglobulin can be obtained by using a cation exchange resin and an ultrafiltration module.
Claims (3)
レンニン等の酵素処理で得た乳清に、エチレンジアミン
四酢酸の濃度0.5-10mM、又はグリシンの濃度が50-500mM
になるように加え、クエン酸ナトリウム溶液で乳清のpH
を6.0-6.5に調整し、これに陽イオン交換樹脂を接触さ
せ、50,000又は100,000ダルトンの分離限界を有する限
外濾過モジユ−ルを用いて、目的とする免疫グロブリン
を得ることを特徴とする免疫グロブリンの製造法。1. A whey obtained by isoelectric point (pH 4.5-4.6) precipitation of casein or enzymatic treatment with rennin, etc., has a concentration of ethylenediaminetetraacetic acid of 0.5-10 mM or a concentration of glycine of 50-500 mM.
And whey pH with sodium citrate solution
Is adjusted to 6.0-6.5, and a cation exchange resin is brought into contact therewith, and the target immunoglobulin is obtained by using an ultrafiltration module having a separation limit of 50,000 or 100,000 daltons. Method for producing globulin.
の分離限界を有する限外濾過モジユ−ルで得た免疫グロ
ブリン画分に陽イオン交換樹脂を接触させ、免疫グロブ
リンの重合体を除去することを特徴とする免疫グロブリ
ン単量体の製造法。2. The immunoglobulin fraction obtained by subjecting the whey obtained in claim 1 to an immunoglobulin fraction obtained by an ultrafiltration module having a separation limit of 100,000 daltons is contacted with a cation exchange resin to remove an immunoglobulin polymer. A method for producing an immunoglobulin monomer, which comprises:
オン交換樹脂とを接触させ、免疫グロブリン重合体を除
去することを特徴とする免疫グロブリン単量体の製造
法。3. A method for producing an immunoglobulin monomer, which comprises contacting the immunoglobulin obtained in claim 1 with a cation exchange resin to remove an immunoglobulin polymer.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP10499194A JPH07285885A (en) | 1994-04-21 | 1994-04-21 | Method for producing immunoglobulin |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP10499194A JPH07285885A (en) | 1994-04-21 | 1994-04-21 | Method for producing immunoglobulin |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH07285885A true JPH07285885A (en) | 1995-10-31 |
Family
ID=14395570
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP10499194A Pending JPH07285885A (en) | 1994-04-21 | 1994-04-21 | Method for producing immunoglobulin |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH07285885A (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1999062936A1 (en) * | 1998-06-01 | 1999-12-09 | Genentech, Inc. | Separation of protein monomers from aggregates by use of ion-exchange chromatography |
| CN1090030C (en) * | 1997-08-06 | 2002-09-04 | 上海科星生物技术有限公司 | Complex cow's milk gamma globulin and the making method thereof |
| US9055752B2 (en) | 2008-11-06 | 2015-06-16 | Intercontinental Great Brands Llc | Shelf-stable concentrated dairy liquids and methods of forming thereof |
| US11490629B2 (en) | 2010-09-08 | 2022-11-08 | Koninklijke Douwe Egberts B.V. | High solids concentrated dairy liquids |
| CN115417929A (en) * | 2022-10-09 | 2022-12-02 | 江苏天美健大自然生物工程有限公司 | Method for extracting bovine colostrum immunoglobulin |
-
1994
- 1994-04-21 JP JP10499194A patent/JPH07285885A/en active Pending
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1090030C (en) * | 1997-08-06 | 2002-09-04 | 上海科星生物技术有限公司 | Complex cow's milk gamma globulin and the making method thereof |
| WO1999062936A1 (en) * | 1998-06-01 | 1999-12-09 | Genentech, Inc. | Separation of protein monomers from aggregates by use of ion-exchange chromatography |
| US6620918B2 (en) | 1998-06-01 | 2003-09-16 | Genentech, Inc. | Separation of polypeptide monomers |
| EP1500661A1 (en) * | 1998-06-01 | 2005-01-26 | Genetech, Inc. | Separation of protein monomers from aggregates by use of ion-exchange chromatography |
| US9055752B2 (en) | 2008-11-06 | 2015-06-16 | Intercontinental Great Brands Llc | Shelf-stable concentrated dairy liquids and methods of forming thereof |
| US11490629B2 (en) | 2010-09-08 | 2022-11-08 | Koninklijke Douwe Egberts B.V. | High solids concentrated dairy liquids |
| CN115417929A (en) * | 2022-10-09 | 2022-12-02 | 江苏天美健大自然生物工程有限公司 | Method for extracting bovine colostrum immunoglobulin |
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