CN1064080C - Industrial production process of velvet deerhorn cell - Google Patents
Industrial production process of velvet deerhorn cell Download PDFInfo
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- CN1064080C CN1064080C CN97111856A CN97111856A CN1064080C CN 1064080 C CN1064080 C CN 1064080C CN 97111856 A CN97111856 A CN 97111856A CN 97111856 A CN97111856 A CN 97111856A CN 1064080 C CN1064080 C CN 1064080C
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- cell
- pilose antler
- growth
- strain
- antler
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Abstract
The present invention relates to a technology for the industrial production of pilose antler cells. Tissues with pilose antler growth are picked so as to be used as raw materials, and anchoring primary pilose antler cells are cultured and are completely adapted to the large scale rotary culture by a rotary bottle. More than 1, 000, 000 pilose antler cells are obtained from each milliliter of culture solution, and the number of cells growing during one period in one culture unit (bottle) can exceed the number of raw antler cells contained in one natural pilose antler.
Description
The present invention relates to the artificial culture method of cell strain, relate to the large-scale industrialization production method of pilose antler cell specifically.
Pilose antler cell is a kind of special mammalian cell of occurring in nature, and the proper splitting speed of growth surpasses cancer cells on the deer body.China people find that pilose antler is medicinal with a long history, clinical efficacy, and health-care effect is distinguished.Modern medicine study has been proved conclusively the active substance and the pharmacological mechanism of pilose antler, extremely praises highly both at home and abroad.
Pilose antler production all depended on since the own beginning wildly hunts or obtains pilose antler in artificial domestic mode, fettered by nature and cultivating condition, the quality good or not great disparity, and production level is limited.
The objective of the invention is to seek a kind of method of large-scale industrial production pilose antler cell, this method is easy and simple to handle, and cost is low, and product should have the identical pharmacological action of natural pilose antler,
The inventor is through repetition test decades, repeatedly select, found out a kind of rolling bottle industrial process of pilose antler cell gradually, this method is taked the starting materials that is organized as of antler growth point, and continuous passage is cultivated and obtained to be suitable for the kind cell strain that extensive rolling bottle is produced in specific cell grown cultures liquid.The present invention has created the new mode of pilose antler material production with biotechnology rolling bottle suitability for industrialized production pilose antler cell, makes its product reach " cured " level fully, and economic development source power is far-reaching.
The object of the present invention is achieved like this:
One, the cultivation of original parent cell line
Take to give birth to fine and soft cell starting materials from the tender antler growth point of children, be characterized as prismatic, the heterocyst of circular different differential periods, under aseptic technique, starting materials is arranged in the Tissue Culture Flask, do adherent culture, add a small amount of (1/10th capacity) cell growth medium, growth media (): prescription
The minimum essential element nutritive liquid of EagleShi (E-MEM) 78%
New-born calve serum 20%
Kantlex 1%
Sodium bicarbonate 1%
In 34 ℃ of cultivations, organize peripheral epitaxy cell from pilose antler, cellular form differs, constantly change growth media according to the growth adaptation situation, inclining earlier original fluid, adds new growth media, individual layer to be grown discards original structure, cell dispersion, doing time pickup kind from primary cell cultivates, cell growth division growth is good, does continuous passage with 0.25% trypsin solution digestion cell dispersion and cultivates, and progressively divides bottle to enlarge according to cell quantity, and excessive to big culturing bottle, temperature is increased to 37 ℃ by 34 ℃.Reach the stagnant phenomenon of giving birth to of appearance about 20 generations when going down to posterity, poor growth, the cell fission phenomenon is vigorous inadequately, has to stop going down to posterity danger, adopt limit and urge living measure, uses growth media (two), the cell continuity is gone down to posterity reached for 30 generations.
Cell growth medium (two) prescription:
E-MEM 90％
New-born calve serum 7%
Cell growth thing (deer blood plasma deer liver leach liquor 2%
1: 10 Eagle liquid)
Kantlex 1%
Sodium bicarbonate is transferred PH7.0-7.2
Two, plant the cultivation of cell strain
The pilose antler parent cell line in 30 generations of going down to posterity of above-mentioned acquisition is inoculated in the cultivation rolling bottle that 1/10th amount growth medias () are housed, the rolling bottle capacity is 3 liters, growth temperature is adjusted to 34-37 ℃, cultivating rotation number and be 6-30 changes/hour, incubation time is 3-7 days, doing time pickup kind then cultivates, culture condition is identical, continuous passage 30-150 generation, reach the growth cycle unanimity, growing way is vigorous, vitality was strong, obtains ideal kind cell strain, and this cell strain has been preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center on June 18th, 1997, preserving number is CGMCC No.0308, this cell strain can be directly used in production or preserve standby in-75 ℃～-195 ℃ very low temperature.
Three, pilose antler cell rolling bottle suitability for industrialized production
Adopt 3 liters, 10 liters or 15 liters to cultivate rolling bottle, the cell growth medium (one) that adds 1/10th capacity, get kind of a cell strain inoculation, inoculum size is about 200,000/ml, under 34-37 ℃ of constant temperature, every 5-8 minute rotation-week (per hour 8-12 changes), cultivate and finished division growth in 24-72 hour, every milliliter can obtain cell 10
6More than, every stage body amasss 2.7-3.00M
3Favourable turn is produced 13,500 ml cells liquid, at 2000M
2The production plant that is equipped with 24 favourable turns can be produced 15,500 liter pilose antler cells per year.
Four, the activity of pilose antler cell, pharmacological action
Activation analysis confirms: contain the corresponding basic thing of clinical efficacy, mainly contain 18 kinds with upper amino acid, 7 kinds of essential amino acids are wherein arranged, 2 kinds of semi-dispensable amino acids: contain phosphatide, polypeptide, especially contain time sulphur purine and spermidine, polyamine substances such as spermine.
The pharmacological testing card: pilose antler cell has significant antifatigue and anti-aging effects, can promote nucleic acid (RNA) and protein synthesis, and the activity of monoamine oxidase of inhibition effect, antioxygenation and the effect of male hormone sample are arranged.
Pharmacological action, medicinal efficacy, active substance are all consistent with natural pilose antler.
Claims (3)
1. pilose antler cell strain, its preserving number is CGMCC NO.0308.
2. the preparation method of a pilose antler cell strain is characterized in that:
(1) takes to give birth to fine and soft cell from the tender antler growth point of children, be arranged in Tissue Culture Flask and do adherent culture, refinement intracellular growth liquid (one), 34 ℃ of cultivation individual layers to be grown, discard original structure, do continuous passage with 0.25% trypsin solution digestion cell dispersion and cultivate, progressively divide bottle to enlarge according to cell quantity, and it is excessive to big culturing bottle, temperature is elevated to 37 ℃ by 34 ℃, and continuous passage was cultured to for 20 generations, when poor growth occurring, use growth media (two), be passaged to for 30 generations;
(2) above-mentioned cell strain is inoculated in the culturing bottle that growth media (one) is housed, temperature is 34-37 ℃, cultivating revolution and be 6-30 changes/hour, incubation time is 3-7 days, continuous passage 30-150 generation, obtains the pilose antler cell strain of claim 1, CGMCC NO.0308;
Wherein the prescription of cell growth medium () is:
E-MEM 78％
New-born calve serum 20%
Kantlex 1%
Soda solution 1%
The prescription of cell growth medium (two) is:
E-MEM 90％
New-born calve serum 7%
Cell growth thing 2%
Kantlex 1%
Sodium bicarbonate is transferred PH7.0-7.2
Described cell growth thing is a deer blood plasma deer liver leach liquor: Eagle liquid is 1: 10.
3. the strain of claim 1 pilose antler cell is used for the purposes of suitability for industrialized production pilose antler cell, it is characterized in that the described pilose antler cell strain of claim 1 is seeded in the cell growth medium of describing in the claim 2 (), under the 34-37 ℃ of constant temperature, rotated a circle in 5-8 minute, cultivated 24-72 hour, and obtained pilose antler cell.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN97111856A CN1064080C (en) | 1997-06-26 | 1997-06-26 | Industrial production process of velvet deerhorn cell |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN97111856A CN1064080C (en) | 1997-06-26 | 1997-06-26 | Industrial production process of velvet deerhorn cell |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1203946A CN1203946A (en) | 1999-01-06 |
| CN1064080C true CN1064080C (en) | 2001-04-04 |
Family
ID=5171918
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN97111856A Expired - Lifetime CN1064080C (en) | 1997-06-26 | 1997-06-26 | Industrial production process of velvet deerhorn cell |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1064080C (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101265463B (en) * | 2007-03-16 | 2013-08-07 | 郑海发 | Culture method for pilose antler cell |
| CN106319637A (en) * | 2016-08-31 | 2017-01-11 | 高伟 | Biotechnology for culturing pilose antler cells and extracting pilose antler active polypeptides in large scale |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN86107813A (en) * | 1985-11-13 | 1987-07-29 | 默里尔多药物公司 | Preparation is as the method for the heterocycle oxazolone class of cardiotonic drug |
-
1997
- 1997-06-26 CN CN97111856A patent/CN1064080C/en not_active Expired - Lifetime
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN86107813A (en) * | 1985-11-13 | 1987-07-29 | 默里尔多药物公司 | Preparation is as the method for the heterocycle oxazolone class of cardiotonic drug |
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| Publication number | Publication date |
|---|---|
| CN1203946A (en) | 1999-01-06 |
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| C53 | Correction of patent for invention or patent application | ||
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Free format text: CORRECT: APPLICANT; FROM: LIANG GUOJIAN; KANG ZHONGYING TO: LIANG GUOJIAN |
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| CP03 | Change of name, title or address |
Address after: 510180 Guangzhou Zhongshan city six road 218-220 Jietai square 17 Applicant after: Liang Guojian Address before: 543004 the Guangxi Zhuang Autonomous Region Wuzhou City Qian Jian Road No. 82 Applicant before: Liang Guojian Applicant before: Kang Zhongying |
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Granted publication date: 20010404 |