CN106167762B - A kind of skin spermatogonium acquisition suit - Google Patents
A kind of skin spermatogonium acquisition suit Download PDFInfo
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- CN106167762B CN106167762B CN201610622081.8A CN201610622081A CN106167762B CN 106167762 B CN106167762 B CN 106167762B CN 201610622081 A CN201610622081 A CN 201610622081A CN 106167762 B CN106167762 B CN 106167762B
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- collecting cassette
- spermatogonium
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M47/00—Means for after-treatment of the produced biomass or of the fermentation or metabolic products, e.g. storage of biomass
- C12M47/04—Cell isolation or sorting
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N1/00—Preservation of bodies of humans or animals, or parts thereof
- A01N1/02—Preservation of living parts
- A01N1/0205—Chemical aspects
- A01N1/021—Preservation or perfusion media, liquids, solids or gases used in the preservation of cells, tissue, organs or bodily fluids
- A01N1/0226—Physiologically active agents, i.e. substances affecting physiological processes of cells and tissue to be preserved, e.g. anti-oxidants or nutrients
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61B—DIAGNOSIS; SURGERY; IDENTIFICATION
- A61B10/00—Other methods or instruments for diagnosis, e.g. instruments for taking a cell sample, for biopsy, for vaccination diagnosis; Sex determination; Ovulation-period determination; Throat striking implements
- A61B10/0096—Casings for storing test samples
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61B—DIAGNOSIS; SURGERY; IDENTIFICATION
- A61B10/00—Other methods or instruments for diagnosis, e.g. instruments for taking a cell sample, for biopsy, for vaccination diagnosis; Sex determination; Ovulation-period determination; Throat striking implements
- A61B10/02—Instruments for taking cell samples or for biopsy
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
- C12M41/12—Means for regulation, monitoring, measurement or control, e.g. flow regulation of temperature
Abstract
The invention discloses the acquisition suits of a kind of skin and/or spermatogonium, include acquisition component in collecting cassette and box；Collecting cassette includes collecting cassette outer cover, collecting cassette inner casing, the first magnet on the inside of collecting cassette outer cover, and second magnet set on collecting cassette inner casing on the outside of corresponding with the first magnet and the fixed pad in the box in collecting cassette inner casing；Acquisition component includes collection tube and the matched double-level-metal ring of collection tube, reagent bottle, capture card and ice chest and the temperature discoloring test paper being fixed on double-level-metal ring in box；Fixed pad is equipped with collection tube card slot, double-level-metal ring card slot, reagent bottle card slot, capture card card slot and the fixed ice chest card slot for padding one end in box in box.Mainly aminoglycoside antibiotics is formed with DMEM culture medium aqueous dissolutions equipped with Cell protective solutions, protection liquid in reagent bottle.This acquisition is easy to use with being set with, and protection liquid fully simulates vivo environment, can significantly improve the survival rate of cell.
Description
Technical field
The present invention relates to a kind of cell collection suits, and in particular to a kind of skin spermatogonium acquisition suit.
Background technology
At present in the clinic in China and cell research experiment, acquiring, preserving and use for a long time for cell is that space-time detaches,
Very harsh to the requirement of external environment after the cells ex vivo of acquisition, the cell particularly between cell collection and long-term preservation turns
During fortune, due to cell have not gone through it is long-term preserve that processing external environment is bad to reduce cell viability, decision acquire successfully and
It is no.While variation of ambient temperature conveniently, cannot be monitored accurately in transhipment transportational process after acquiring, it is not known that be had in transportational process
Without causing cell inactivation because environment temperature is unqualified.Therefore, cell collection is easily and efficiently carried out, the environment created
Cell is transported through, and it is monitored, the clinic and a critically important subject in cell research experiment for being China.
Invention content
The present invention provides an acquisition suits, can easily and efficiently carry out cell collection, and the environment pair created
Cell is transported through, the protection liquid in acquisition suit, it is further provided skin spermatogonium suitable and stablize Conservation environment,
The cell viability after skin spermatogonium preserves can be significantly improved.Specific technical solution is as follows：
A kind of skin spermatogonium acquisition suit, includes acquisition component in collecting cassette and box；
The collecting cassette include collecting cassette outer cover, collecting cassette inner casing, the first magnet on the inside of collecting cassette outer cover, with the
One magnet correspondence and the second magnet on the outside of collecting cassette inner casing and the fixed pad in the box in collecting cassette inner casing；
Acquisition component includes collection tube and the matched double-level-metal ring of collection tube, reagent bottle, capture card and ice in the box
Box, the temperature discoloring test paper being fixed on double-level-metal ring；
In the box fixed pad equipped with collection tube card slot, double-level-metal ring card slot, reagent bottle card slot, capture card card slot and
The ice chest card slot of fixed pad one end in box.
Preferably, the collecting cassette outer cover and collecting cassette inner casing material are PC plastic.
Preferably, the collection tube is PC test tubes with a scale and pipe cap.
Preferably, the double-level-metal ring material is aluminium.
Preferably, also to include bar code a set of for acquisition component in the box.
Preferably, acquisition component also includes acquisition suit operation instruction portion in the box.
Preferably, equipped with Cell protective solutions in the reagent bottle, the protection liquid is mainly by aminoglycoside antibiotics
It is formed with DMEM culture medium aqueous dissolutions, aminoglycoside antibiotics described in every milliliter of DMEM culture mediums aqueous dissolution
50-200 units, a concentration of 10-25g/L of DMEM culture mediums, the aminoglycoside resist in the DMEM culture mediums aqueous solution
Raw element is one or more in gentamicin, kanamycins, amikacin or tobramycin.
Preferably, the protection liquid further includes oxyhemoglobin, every milliliter of DMEM culture mediums aqueous dissolution institute
State oxyhemoglobin 400-1000 μ g.
Preferably, the protection liquid further includes one or more, every milliliter of DMEM culture in vitamin E or glutathione
0.25 μm of ol of 0.25 μm of ol of base aqueous dissolution vitamin E and glutathione.
Preferably, the protection liquid further includes Tween 80, palmitamide, potassium alginate and Hamposyl L, every milliliter
The DMEM culture mediums aqueous solution dissolves 0.5 μ L of Tween 80,60 μ g of palmitamide, 30 μ g of potassium alginate, lauroyl flesh ammonia respectively
150 μ g of acid.
Preferably, the protection liquid further includes citric acid, pyruvic acid, fumaric acid and acetyl coenzyme A, described in every milliliter
DMEM culture mediums aqueous solution dissolves citric acid 1-10 μ g, pyruvic acid 50-80 μ g, fumaric acid 10-20 μ g and acetylcoenzyme respectively
A80-150μg。
The beneficial effects of the invention are as follows：Acquisition suit provided by the invention is easy to use, former convenient for Skin Cell or essence
The storage and transport of cell, and the protection liquid in reagent bottle fully simulates vivo environment, is more suitable for Skin Cell and essence is former
The existence of cell can significantly improve the survival rate of more than cell.
Description of the drawings
Fig. 1 is the bottom view of 1 collecting cassette outer cover of embodiment；
Fig. 2 is the acquisition suit vertical view that embodiment 1 opens collecting cassette outer cover；
In figure, 1 collecting cassette outer cover, 2 collecting cassette inner casings, 3 first magnet, 4 second magnet, fixed pad, 11 collection tubes in 5 boxes
Card slot, 12 double-level-metal ring card slots, 13 reagent bottle card slots, 14 capture card card slots, 15 ice chest card slots.
Specific embodiment
Embodiment 1
A kind of skin spermatogonium acquisition suit, includes acquisition component in collecting cassette and box；
The collecting cassette include collecting cassette outer cover 1, collecting cassette inner casing 2, set on the inside of collecting cassette outer cover 1 the first magnet 3,
It is corresponding with the first magnet 3 and fix pad 5 in the second magnet 4 and box on the outside of collecting cassette inner casing；
Acquisition component includes collection tube and the matched double-level-metal ring of collection tube, reagent bottle, capture card, ice in the box
Box and the temperature discoloring test paper being fixed on double-level-metal ring；
Pad 5 is fixed in the box equipped with collection tube card slot 11, double-level-metal ring card slot 12, reagent bottle card slot 13, acquisition
Card card slot 14 and the fixed ice chest card slot 15 for padding 5 one end in box.
The acquisition suit collecting cassette outer cover of front opening skin spermatogonium is acquired, ready ice chest is taken to be put into ice chest card
Slot.
Collection tube is then taken out, then the skin spermatogonium of acquisition is put into collection tube, often pipe 25ml.It will be in reagent bottle
Protection liquid press 1:1 ratio is poured into collection tube, tightens collection tube lid.Then it is marked on collection tube with marking pen.
Then inspection and the matched temperature discoloring test paper being fixed in double-level-metal ring of collection tube, will after confirming normally
Being mounted on collection tube with the matched double-level-metal ring of collection tube for temperature discoloring test paper is fixed with, becket is close to collection tube.
When collection tube needs temporarily to preserve, 4 DEG C of refrigerators are put into, while ensure that collection tube pipe lid is tightened, collection tube is upright, steady
It is fixed, it can be preserved within 12 hours.
Then donor data and acquisition information are filled in detail on capture card.Before acquisition suit handing-over transport, acquisition is replaced
Cover ice chest card slot in ice chest, by collection tube, fill in complete capture card by origin-location be put into acquisition suit in, cover acquisition
Box outer cover ensures to be set on the first magnet on the inside of collecting cassette outer cover and the second magnetic on the outside of inner casing corresponding with the first magnet
Iron attracts each other.
Transport is completed, and opens the acquisition suit of skin spermatogonium, is checked and collection tube is matched is fixed on becket
Temperature discoloring test paper, confirm it is normal after, complete handing-over, measure carried out after cell viability meets the requirements subsequent long-term preservation or
Person is proliferated.If it is undesirable for acquisition to measure cell viability higher than the requirement temperature to environment for the test paper displays temperature that changes colour
Failure needs to abandon the skin spermatogonium of acquisition.
The acquisition suit of the skin spermatogonium of the present embodiment, it is easy to use, it is cheap.In collecting cassette outer cover
First magnet and the second magnet on the outside of inner casing corresponding with the first magnet attract each other, and reduce and are adopted in transportational process of knowing clearly
The probability that collection box outer cover is opened due to shaking or other accidents.Ice in ice chest provides the low temperature environment in collecting cassette, ice chest
Card slot is located at one end rather than bottom of fixed pad in box, if haulage time is longer, in the case that ice is inadequate, can not take
Go out in box to carry out the replacement of ice chest in the case of fixed pad, can preferably ensure low temperature environment.Temperature discoloring test paper monitoring fortune
Temperature change during defeated.Shield temperature discoloring test paper is easily received water or the pollution of other impurity and fails, and is not suitable for directly
It is attached to monitoring problem on collection tube.Common plastic protective layer can be with insulation blocking temperature discoloring test paper, but also completely cuts off simultaneously
Heat, thermal conductivity is bad, the temperature of temperature discoloring test paper and the temperature of collection tube have a certain distance.The bilayer of the present embodiment
Becket sandwiches protection temperature discoloring test paper, and isolation makes it not by water or other pollutions, while be close to collection tube, preferably transmits
Heat so that the closer temperature with collection tube of temperature discoloring test paper temperature.
The acquisition suit internal temperature measure of the change of skin spermatogonium, first takes the ice chest prepared, is put into ice chest card slot,
Thermometer is put into, after being pre-chilled 30 minutes, the temperature of thermometer is observed through transparent collecting cassette and records every 30 minutes records one
Secondary temperature.Acquisition suit internal temperature is consistently higher than 4 degrees Celsius in 12 hours.
Acquisition suit inner skin 12 hours Dynamic Curves of spermatogonium measure, mtt assay measure cell viability (《Wang Zhirong is high
Fine jade, Ma Chuanxin, Li Yan girl .MTT methods measure Escherichia coli viable count experimental study, ACTA Scientiae Circumstantiae, 2011,31 (12)：
2642-1650.), sampling in every 30 minutes is primary, using the vigor of the first sub-sampling as 100%.
Embodiment 2
Difference lies in it is a set of also to include bar code to the present embodiment for acquisition component in the box with embodiment 1.
In use, directly bar code is attached on collection tube, the time can be saved in this way, accelerates speed, it is thus also avoided that note
The risk that number pen label is accidentally wiped in transit.
Embodiment 3
Difference lies in acquisition component also includes acquisition suit operation instruction to the present embodiment in the box with embodiment 1
It is a.
For the people that the acquisition for not using this skin spermatogonium in the past is set with, can be carried out according to operation instruction ripe
Experienced operation.
Embodiment 4
Difference lies in the collection tube is PC test tubes with a scale and pipe cap to the present embodiment, described double with embodiment 1
Layer becket material is aluminium, and Cell protective solutions are housed in the reagent bottle.
Embodiment 5 protects the preparation, use and cell viability of liquid to measure
Protect liquid 1
A kind of skin stem spermatogonium protects liquid, which is mainly that gentamicin is molten with DMEM culture medium aqueous solutions
Solve, every milliliter of 100 unit of DMEM culture mediums aqueous dissolution gentamicin, the DMEM culture mediums aqueous solution it is dense
It spends for 16.9g/L.
Protect liquid 2
A kind of skin stem spermatogonium protects liquid, which is mainly that kanamycins is molten with DMEM culture medium aqueous solutions
Solve, every milliliter of 100 unit of DMEM culture mediums aqueous dissolution kanamycins, the DMEM culture mediums aqueous solution it is dense
It spends for 16.9g/L.
Protect liquid 3
A kind of skin stem spermatogonium protects liquid, which is mainly that amikacin is molten with DMEM culture medium aqueous solutions
Solve, every milliliter of 100 unit of DMEM culture mediums aqueous dissolution amikacin, the DMEM culture mediums aqueous solution it is dense
It spends for 16.9g/L.
Protect liquid 4
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin, kanamycins and tobramycin
Formed with DMEM culture medium aqueous dissolutions, every milliliter of DMEM culture mediums aqueous solution dissolve respectively 100 unit of gentamicin,
20 unit of kanamycins, 20 unit of tobramycin, a concentration of 16.9g/L of the DMEM culture mediums aqueous solution.
Protect liquid 5
A kind of skin stem spermatogonium protects liquid, which is mainly that tobramycin is molten with DMEM culture medium aqueous solutions
Solve, every milliliter of 100 unit of DMEM culture mediums aqueous dissolution tobramycin, the DMEM culture mediums aqueous solution it is dense
It spends for 10.0g/L.
Protect liquid 6
A kind of skin stem spermatogonium protects liquid, which is mainly that gentamicin is molten with DMEM culture medium aqueous solutions
Solve, every milliliter of 100 unit of DMEM culture mediums aqueous dissolution gentamicin, the DMEM culture mediums aqueous solution it is dense
It spends for 25.0g/L.
Protect liquid 7
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin and oxyhemoglobin DMEM
Culture medium aqueous dissolution forms, and every milliliter of DMEM culture mediums aqueous solution dissolves 100 unit of gentamicin, oxygenated blood respectively
Lactoferrin 400 μ g, a concentration of 16.9g/L of the DMEM culture mediums aqueous solution.
Protect liquid 8
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin and oxyhemoglobin DMEM
Culture medium aqueous dissolution forms, and every milliliter of DMEM culture mediums aqueous solution dissolves 100 unit of gentamicin, oxygenated blood respectively
Lactoferrin 600 μ g, a concentration of 16.9g/L of the DMEM culture mediums aqueous solution.
Protect liquid 9
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin and oxyhemoglobin DMEM
Culture medium aqueous dissolution forms, and every milliliter of DMEM culture mediums aqueous solution dissolves 100 unit of gentamicin, oxygenated blood respectively
Lactoferrin 1000 μ g, a concentration of 16.9g/L of the DMEM culture mediums aqueous solution.
Protect liquid 10
A kind of skin stem spermatogonium protects liquid, which mainly cultivates gentamicin and vitamin E with DMEM
Base aqueous dissolution forms, and every milliliter of DMEM culture mediums aqueous solution dissolves 100 unit of gentamicin, vitamin E 0.1 respectively
μm ol, a concentration of 16.9g/L of the DMEM culture mediums aqueous solution.
Protect liquid 11
A kind of skin stem spermatogonium protects liquid, which mainly uses gentamicin, vitamin E and glutathione
DMEM culture medium aqueous dissolutions form, and every milliliter of DMEM culture mediums aqueous solution dissolves 100 unit of gentamicin, dimension respectively
E0.3 μm of ol of raw element, glutathione 0.3 μm of ol, a concentration of 16.9g/L of the DMEM culture mediums aqueous solution.
Protect liquid 12
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin, oxyhemoglobin, vitamin
E and glutathione are formed with DMEM culture medium aqueous dissolutions, and it is big that every milliliter of DMEM culture mediums aqueous solution dissolves celebrating respectively
0.25 μm of 100 unit of mycin, 600 μ g of oxyhemoglobin, 0.25 μm of ol of vitamin E, glutathione ol, the DMEM culture mediums
A concentration of 16.9g/L of aqueous solution.
Protect liquid 13
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin, oxyhemoglobin, vitamin
C and vitamin E are formed with DMEM culture medium aqueous dissolutions, and it is big mould that every milliliter of DMEM culture mediums aqueous solution dissolves celebrating respectively
0.5 μm of plain 100 units, 600 μ g of oxyhemoglobin, 0.5 μm of ol of vitamin C, vitamin E ol, the DMEM culture mediums are water-soluble
A concentration of 16.9g/L of liquid.
Protect liquid 14
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin, Tween 80, palmitamide, sea
Potassium alginate and Hamposyl L are formed with DMEM culture medium aqueous dissolutions, every milliliter of DMEM culture mediums aqueous solution difference
100 unit of gentamicin, 0.5 μ g of Tween 80,80 μ g of palmitamide, 50 μ g of potassium alginate, 100 μ g of Hamposyl L are dissolved,
A concentration of 16.9g/L of the DMEM culture mediums aqueous solution.Protect liquid 15
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin, oxyhemoglobin, tween
80th, palmitamide, potassium alginate and Hamposyl L are formed with DMEM culture medium aqueous dissolutions, every milliliter of DMEM training
It supports base aqueous solution and dissolves 100 unit of gentamicin, 600 μ g of oxyhemoglobin, 1.0 μ g of Tween 80,20 μ of palmitamide respectively
G, 10 μ g of potassium alginate, Hamposyl L 200 μ g, a concentration of 16.9g/L of the DMEM culture mediums aqueous solution.
Protect liquid 16
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin, oxyhemoglobin, vitamin
E, glutathione, Tween 80, palmitamide, potassium alginate and Hamposyl L are formed with DMEM culture medium aqueous dissolutions,
Every milliliter of DMEM culture mediums aqueous solution dissolves 100 unit of gentamicin, oxyhemoglobin 600mg, vitamin respectively
E0.25 μm of ol, 0.25 μm of ol of glutathione, 0.5 μ g of Tween 80,60 μ g of palmitamide, 30 μ g of potassium alginate, lauroyl flesh ammonia
150 μ g, a concentration of 16.9g/L of the DMEM culture mediums aqueous solution of acid.
Protect liquid 17
A kind of skin stem spermatogonium protects liquid, which is mainly by gentamicin, citric acid, pyruvic acid, prolongs recklessly
Rope acid and acetyl coenzyme A are formed with DMEM culture medium aqueous dissolutions, every milliliter of DMEM culture mediums aqueous dissolution difference
100 unit of gentamicin, 10 μ g of citric acid, 80 μ g of 80 μ g of pyruvic acid, 10 μ g of fumaric acid and acetyl coenzyme A, the DMEM trainings
Support a concentration of 16.9g/L of base aqueous solution.
Protect liquid 18
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin, oxyhemoglobin, vitamin
E, glutathione, citric acid, pyruvic acid, fumaric acid and acetyl coenzyme A are formed with DMEM culture medium aqueous dissolutions, every milliliter
The DMEM culture mediums aqueous solution dissolves 100 unit of gentamicin, 600 μ g of oxyhemoglobin, 0.25 μ of vitamin E respectively
Mol, 0.25 μm of ol of glutathione, 1 μ g of citric acid, 50 μ g of pyruvic acid, 20 μ g of fumaric acid and acetyl coenzyme A 150 μ g, it is described
A concentration of 16.9g/L of DMEM culture medium aqueous solutions.
Protect liquid 19
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin, oxyhemoglobin, vitamin
E, glutathione, Tween 80, palmitamide, potassium alginate, Hamposyl L, citric acid, pyruvic acid, fumaric acid and acetyl
Coacetylase is formed with DMEM culture medium aqueous dissolutions, and every milliliter of DMEM culture mediums aqueous solution dissolves gentamicin 100 respectively
Unit, 600 μ g of oxyhemoglobin, 0.25 μm of ol of vitamin E, 0.25 μm of ol of glutathione, 0.5 μ g of Tween 80, palmitamide
60 μ g, 30 μ g of potassium alginate, 150 μ g of Hamposyl L, 4 μ g of citric acid, 60 μ g of pyruvic acid, 15 μ g of fumaric acid and acetyl are auxiliary
Enzyme A 120 μ g, a concentration of 16.9g/L of the DMEM culture mediums aqueous solution.
Comparison liquid 1
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by penicillin DMEM culture medium aqueous dissolutions
Form, every milliliter of 100 unit of DMEM culture mediums aqueous dissolution penicillin, the DMEM culture mediums aqueous solution it is a concentration of
16.9g/L。
Comparison liquid 2
A kind of skin stem spermatogonium protects liquid, which mainly cultivates gentamicin and transferrins with DMEM
Base aqueous dissolution forms, and every milliliter of DMEM culture mediums aqueous solution dissolves 100 unit of gentamicin, transferrins respectively
400 μ g, a concentration of 16.9g/L of the DMEM culture mediums aqueous solution.
Comparison liquid 3
A kind of skin stem spermatogonium protects liquid, which mainly uses gentamicin, vitamin E and glutathione
DMEM culture medium aqueous dissolutions form, and every milliliter of DMEM culture mediums aqueous solution dissolves 100 unit of gentamicin, dimension respectively
E0.1 μm of ol of raw element, glutathione 0.9 μm of ol, a concentration of 16.9g/L of the DMEM culture mediums aqueous solution.
Comparison liquid 4
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin, sorbester p17, palmitamide, sea
Potassium alginate and Hamposyl L are formed with DMEM culture medium aqueous dissolutions, every milliliter of DMEM culture mediums aqueous solution difference
100 unit of gentamicin, 0.5 μ g of sorbester p17,80 μ g of palmitamide, 50 μ g of potassium citrate, 100 μ g of Hamposyl L are dissolved,
A concentration of 16.9g/L of the DMEM culture mediums aqueous solution.Comparison liquid 5
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin, Tween 80, palmitamide and sea
Potassium alginate is formed with DMEM culture medium aqueous dissolutions, and every milliliter of DMEM culture mediums aqueous solution dissolves gentamicin respectively
100 units, 0.5 μ g of Tween 80,80 μ g of palmitamide, 50 μ g of potassium alginate, the DMEM culture mediums aqueous solution it is a concentration of
16.9g/L。
Comparison liquid 6
A kind of skin stem spermatogonium protects liquid, the protection liquid mainly by gentamicin, potassium citrate, Sodium Pyruvate,
Fumaric acid and acetyl coenzyme A are formed with DMEM culture medium aqueous dissolutions, every milliliter of DMEM culture mediums aqueous solution difference
Dissolve 100 unit of gentamicin, 10 μ g of potassium citrate, 80 μ g of 80 μ g of Sodium Pyruvate, 10 μ g of fumaric acid and acetyl coenzyme A, institute
State a concentration of 16.9g/L of DMEM culture medium aqueous solutions.
Comparison liquid 7
A kind of skin stem spermatogonium protects liquid, and the protection liquid is mainly by gentamicin, citric acid, pyruvic acid and Yan Hu
Rope acid is formed with DMEM culture medium aqueous dissolutions, and every milliliter of DMEM culture mediums aqueous solution dissolves gentamicin 100 respectively
Unit, 10 μ g of citric acid, 80 μ g of pyruvic acid and fumaric acid 10 μ g, a concentration of 16.9g/L of the DMEM culture mediums aqueous solution.
Test example 1
The sterilizing protection liquid for being stored in 4 DEG C of refrigerators is taken, with the Skin Cell or spermatogonium that just acquire by 1:1 volume ratio
It is uniformly mixed, is placed in 4 DEG C of environment, different time points, sampling should be carried out, cell viability is measured with mtt assay, with the first sub-sampling
Vigor be 100%.
The main influence for investigating aminoglycoside antibiotics and DMEM concentration of this group experiment, is preserved respectively with comparison liquid 1
Skin Cell and spermatogonium are 1 group of control, to protect Skin Cell and spermatogonium that liquid 2-6 preserves respectively to test 1-5
The Skin Cell and spermatogonium of group, each experimental group and control group enter test procedure after 24h respectively, adjustment Skin Cell and
The density of spermatogonium is 1 × 106cells/mL.By cell suspension：0.4% trypan blue=3:1(v:V) abundant mixing, takes
20 μ L cell suspensions are added in cell counting board, detect each experimental group with Countstar cell counters and control group protects liquid
Endepidermis stem cell and the motility rate of stem spermatogonium, the results are shown in Table 1.
1 test example 1 of table respectively protects the Cell viability result of liquid and comparison liquid
As shown in Table 1, the Skin Cell and the Cell viability of spermatogonium that protection liquid 1-6 is preserved respectively are than using control
The Skin Cell and the Cell viability of spermatogonium that the protection liquid that 1 group of liquid uses preserves respectively are high；The protection liquid that protection liquid 4 uses
The Skin Cell and the Cell viability of spermatogonium preserved respectively is protected respectively than the protection liquid that protection liquid 1-3,5-6 is used to use
The Skin Cell and the Cell viability of spermatogonium deposited are high.
It follows that the aminoglycoside antibiotics in protection liquid provided by the invention has changed the antibiotic such as penicillin into
Afterwards, the motility rate of Skin Cell or spermatogonium significantly reduces, and aminoglycoside antibiotics is using gentamicin, kanamycins
After the mixture of tobramycin, the Skin Cell of preservation and the Cell viability higher of spermatogonium.
Test example 2
The main influence for investigating oxyhemoglobin of this group experiment, the Skin Cell preserved respectively with comparison liquid 2 and essence are former
Cell is 2 groups of control, to protect Skin Cell and spermatogonium that liquid 1,7 and 8 preserves respectively to test 7-9 groups, detection process
Such as test example 1, the testing result such as table 2 after preserving for 24 hours.
2 test example 2 of table respectively protects the Cell viability result of liquid and comparison liquid
As shown in Table 2, the Skin Cell and the Cell viability of spermatogonium that protection liquid 7,8 preserves respectively are than using control
The Skin Cell and the Cell viability of spermatogonium that the protection liquid that 2 groups of liquid uses preserves respectively are high；The skin that protection liquid 8 preserves respectively
The Cell viability of skin cell and spermatogonium is than using the cell of the Skin Cell that liquid 7 is protected to preserve respectively and spermatogonium to live
Rate is high.
It follows that after the oxyhemoglobin in protection liquid provided by the invention has changed transferrins into, Skin Cell
Or the motility rate of spermatogonium significantly reduces, and and during a concentration of 600 μ g/mL of oxyhemoglobin, skin that protection liquid preserves respectively
The high higher of the Cell viability of skin cell and spermatogonium.
Test example 3
The main influence for investigating antioxidant of this group experiment, the Skin Cell and spermatogonium preserved respectively with comparison liquid 3
For control group, protection liquid 7,10-12 are experimental group, and detection process such as test example 1 preserves the testing result such as table 2 after 48h.
3 test example 3 of table respectively protects the Cell viability result of liquid and comparison liquid
As shown in Table 3, the Skin Cell and the Cell viability of spermatogonium that protection liquid 10-12 is preserved respectively are protected than using
The Skin Cell and the Cell viability of spermatogonium that the protection liquid that shield liquid 7,3 groups of comparison liquid use preserves respectively are high；
The Skin Cell and the Cell viability of spermatogonium that protection liquid 12 preserves respectively are than using protection liquid 10,11 to distinguish
The Skin Cell of preservation and the Cell viability of spermatogonium are high.
It follows that the antioxidant in protection liquid provided by the invention can significantly improve Skin Cell or spermatogonium
Motility rate.
Test example 4
Using the comparison liquid 4-7 Skin Cells preserved respectively and spermatogonium as control group, protection liquid 4, protection liquid 7, protection
Liquid 10, protection liquid 14,16, protection liquid 17 and protection liquid 19 are Skin Cell and the essence original of experimental group, each experimental group and control group
Cell enters test procedure for 24 hours, after 48h, 96h, 240h, 480h respectively, and the density for adjusting Skin Cell and spermatogonium is equal
For 1 × 106cells/mL.By cell suspension：0.4% trypan blue=3:1(v:V) abundant mixing takes 20 μ L cell suspensions to add in
In cell counting board, each experimental group and control group protection liquid endepidermis stem cell and essence are detected with Countstar cell counters
The motility rate of former stem cell, the results are shown in Table 4.
4 test example 4 of table respectively protects the Cell viability result of liquid and comparison liquid
As shown in Table 4, the Skin Cell and the cell of spermatogonium that protection liquid 14,16-17 and protection liquid 19 preserve respectively
Motility rate is than using protection liquid 1, protection liquid 7 and protecting the Skin Cells that preserve respectively of 10 comparison liquid 4-7 of liquid and spermatogonium
Cell viability is high；Protection liquid 17 and the time of the Skin Cell that preserves respectively of liquid 19 and spermatogonium is protected than using protection liquid
14 and 16 holding time is long.
It follows that Tween 80, palmitamide, potassium alginate and Hamposyl L in protection liquid provided by the invention
Composition can significantly improve the motility rate of Skin Cell or spermatogonium, citric acid, pyruvic acid, fumaric acid and acetyl coenzyme A
Composition can significantly improve holding time of the protection liquid to Skin Cell or spermatogonium.
Claims (6)
1. a kind of acquisition suit of epidermal stem cells and stem spermatogonium, it is characterised in that：Include in collecting cassette and box and acquire
Component；
The collecting cassette includes collecting cassette outer cover (1), collecting cassette inner casing (2), the first magnet on the inside of collecting cassette outer cover (1)
(3), corresponding with the first magnet (3) and the second magnet (4) on the outside of the collecting cassette inner casing (2) and set on collecting cassette inner casing (2)
Fixed pad (5) in interior box；
In the box acquisition component include collection tube, with the matched double-level-metal ring of collection tube, reagent bottle, capture card, ice chest and
The temperature discoloring test paper being fixed on double-level-metal ring；
In the box fixed pad (5) be equipped with collection tube card slot (11), double-level-metal ring card slot (12), reagent bottle card slot (13),
Capture card card slot (14) and the fixed ice chest card slot (15) for padding (5) one end in box；Cytoprotection is housed in the reagent bottle
Liquid, the protection liquid mainly forms aminoglycoside antibiotics with DMEM culture medium aqueous dissolutions, described in every milliliter
Aminoglycoside antibiotics 50-200 units described in DMEM culture medium aqueous dissolutions, DMEM in the DMEM culture mediums aqueous solution
A concentration of 10-25g/L of culture medium, the aminoglycoside antibiotics are selected from gentamicin, kanamycins, amikacin or appropriate
It is one or more in Obramycin；The protection liquid further includes oxyhemoglobin, every milliliter of DMEM culture mediums aqueous solution
Dissolve the oxyhemoglobin 400-1000 μ g；The protection liquid further includes vitamin E and glutathione, every milliliter of DMEM training
Support 0.25 μm of ol of 0.25 μm of ol of base aqueous dissolution vitamin E and glutathione；The protection liquid further includes Tween 80, palmityl
Amine, potassium alginate and Hamposyl L, every milliliter of DMEM culture mediums aqueous solution dissolve 0.5 μ L of Tween 80, palm respectively
60 μ g of amide, 30 μ g of potassium alginate, 150 μ g of Hamposyl L.
2. the acquisition suit of epidermal stem cells as described in claim 1 and stem spermatogonium, which is characterized in that the acquisition
Box outer cover (1) and collecting cassette inner casing (2) material are PC plastic.
3. the acquisition suit of epidermal stem cells as described in claim 1 and stem spermatogonium, which is characterized in that the acquisition
It manages as with a scale and pipe cap PC test tubes.
4. the acquisition suit of epidermal stem cells as described in claim 1 and stem spermatogonium, which is characterized in that the double-deck gold
It is aluminium to belong to ring material.
5. the acquisition suit of epidermal stem cells as described in claim 1 and stem spermatogonium, which is characterized in that adopted in the box
It is a set of that collection component also includes bar code.
6. the acquisition suit of epidermal stem cells as described in claim 1 and stem spermatogonium, which is characterized in that adopted in the box
It is a that collection component also includes acquisition suit operation instruction.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610622081.8A CN106167762B (en) | 2016-08-01 | 2016-08-01 | A kind of skin spermatogonium acquisition suit |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610622081.8A CN106167762B (en) | 2016-08-01 | 2016-08-01 | A kind of skin spermatogonium acquisition suit |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN106167762A CN106167762A (en) | 2016-11-30 |
| CN106167762B true CN106167762B (en) | 2018-06-22 |
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| Application Number | Title | Priority Date | Filing Date |
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| CN201610622081.8A Active CN106167762B (en) | 2016-08-01 | 2016-08-01 | A kind of skin spermatogonium acquisition suit |
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| CN202015626U (en) * | 2011-03-15 | 2011-10-26 | 高涵 | Portable kit for high school chemistry experiments |
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| CN203283599U (en) * | 2012-12-05 | 2013-11-13 | 江苏和泽生物科技有限公司 | Placenta collection set |
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| CN105062959A (en) * | 2015-09-20 | 2015-11-18 | 领航干细胞再生医学工程有限公司 | Isolated culture method of human amnia mesenchymal stem cells |
| CN105532641A (en) * | 2015-12-28 | 2016-05-04 | 贵州泰邦生物制品有限公司 | Placenta preserving fluid and use method thereof |
| CN206014907U (en) * | 2016-08-01 | 2017-03-15 | 北京世纪劲得生物技术有限公司 | A kind of collection suit of skin spermatogonium |
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2016
- 2016-08-01 CN CN201610622081.8A patent/CN106167762B/en active Active
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN202015626U (en) * | 2011-03-15 | 2011-10-26 | 高涵 | Portable kit for high school chemistry experiments |
| CN202738683U (en) * | 2012-07-25 | 2013-02-20 | 臻景生物技术（上海）有限公司 | Tissue cryopreservation liquid kit |
| CN203283599U (en) * | 2012-12-05 | 2013-11-13 | 江苏和泽生物科技有限公司 | Placenta collection set |
| CN204039387U (en) * | 2014-09-17 | 2014-12-24 | 黑龙江天晴干细胞有限公司 | A kind of acquisition process test kit for obtaining stem cell in tooth |
| CN105062959A (en) * | 2015-09-20 | 2015-11-18 | 领航干细胞再生医学工程有限公司 | Isolated culture method of human amnia mesenchymal stem cells |
| CN105532641A (en) * | 2015-12-28 | 2016-05-04 | 贵州泰邦生物制品有限公司 | Placenta preserving fluid and use method thereof |
| CN206014907U (en) * | 2016-08-01 | 2017-03-15 | 北京世纪劲得生物技术有限公司 | A kind of collection suit of skin spermatogonium |
Also Published As
| Publication number | Publication date |
|---|---|
| CN106167762A (en) | 2016-11-30 |
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