Disinfectant for primary cell culture of animals and preparation method of disinfectant

A technology of animal cells and disinfectants, applied in botany equipment and methods, chemicals for biological control, disinfectants, etc., can solve the problems of not being able to prevent bacterial and mold pollution at the same time, narrow antibacterial spectrum, etc., and achieve high efficiency Disinfection, effect of broad antimicrobial spectrum

Inactive Publication Date: 2018-06-22
CHINA JILIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

This new chemical has two main technical benefits: it does kill many types of germs while still being effective at killing them; it also prevents certain kinds from growing on various materials that are important for their healthy functioning (such as milk).

Problems solved by technology

This patents discuss various methods for controllably reducing harmful organism levels within laboratories' facilities while minimizing damage caused by these agents. However, current techniques require manual identification and adjustments based upon factors like sensitivity to each agent over several hours before they become effective. Additionally, there exist concerns about accurate determining the type of microscopically intact colonized specimen being treated because conventional methods often result in incomplete removal of all infected areas even if complete elimination occurs.

Method used

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  • Disinfectant for primary cell culture of animals and preparation method of disinfectant
  • Disinfectant for primary cell culture of animals and preparation method of disinfectant
  • Disinfectant for primary cell culture of animals and preparation method of disinfectant

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Embodiment 1: the preparation of compound disinfectant

[0051] Accurately weigh 25 mg of amphotericin B, add 20.5 mg of sodium deoxycholate, add 10 ml of sterile water for injection, add 2.0 ml of 0.1 mol / L sodium hydroxide solution dropwise until dissolved (pH value is about 11.5), and then add 0.1 mol / L hydrochloric acid solution 1.8ml (90% of sodium hydroxide consumption) is solution 1. Weigh 1,000,000 U of penicillin, 1 g of streptomycin sulfate, and 0.8 g of gentamycin sulfate in 80 ml of PBS buffer and dissolve to form solution 2. After mixing solution 1 and solution 2, dilute to 100ml with PBS buffer, filter and sterilize with a 0.22um sterile microporous membrane, then pack 5ml into vials, and store at -20°C in the dark. The above-mentioned antibiotic stock solution is a 100-fold stock solution, which should be diluted with culture medium or corresponding buffer before use.

Embodiment 2

[0052] Example 2: Primary culture of rat vascular smooth muscle tissue cells in vitro

[0053] 1 Take a rat of about 200g-250g, anesthetize it with ether and fix it on the operating table.

[0054] 2 Disinfect the skin with 75% ethanol solution, cut the skin with ophthalmic scissors, separate and take out the thoracic aorta with new sterile ophthalmic forceps and ophthalmic scissors, and then put it into a petri dish filled with pre-cooled disinfection buffer PBS; or separate The blood vessels that come down are put into the culture dish of pre-iced DMEM, and quickly transferred to the ultra-clean workbench of the cell culture room.

[0055] 3. Separate the vascular media in the ultra-clean bench, use ophthalmic tweezers to remove blood clots and adipose tissue outside the blood vessels, then use ophthalmic scissors to cut the blood vessels, and use two ophthalmic curved tweezers to scrape off the vascular intima. Then scrape the vascular media. Use one curved forceps to pre...

Embodiment 3

[0062] Example 3: Primary Cell Culture of Female Rat Urethral Tissue

[0063] 1. Feminine rats of about 200g-250g were killed by decapitation, and the urethral tissues were dissected and separated.

[0064] 2 Separate the rat urethral tissue pieces into about 1mm in pre-cooled PBS buffer 3 of small pieces.

[0065] 3 will be 1mm 3 Put the tissue pieces into 10ml of pre-cooled PBS buffer containing compound disinfectant for 20min, then discard the liquid.

[0066] 4 Rinse the tissue block 3 times with pre-cooled PBS buffer.

[0067] 5 Use a sterile Pasteur pipette to put the tissue block into a 24 culture plate, add a small amount of DMEM medium containing a compound disinfectant, and place it in a 5% carbon dioxide incubator at 37°C for about 4 hours.

[0068] 6. Add medium to a total volume of about 1.5ml medium, and place in 37°C containing 5% carbon dioxide for primary cell culture.

[0069] 7According to the growth of the cells, after about 12 days, carry out cell exp...

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Abstract

The invention belongs to the technical field of animal cell engineering and particularly relates to a disinfectant for in vitro cell culture of animals and a preparation method of the disinfectant. The compound disinfectant is mainly prepared from penicillin, streptomycin sulfate, gentamicin sulfate and amphotericin B. Under the condition that cell survival is not affected, the compound disinfectant provided by the invention has a comprehensive antimicrobial spectrum, can simultaneously inhibit the growth of most of Gram-positive bacteria, Gram-negative bacteria and molds, realizes primary culture and tissue sample disinfection of animal cells, and prevents microbial contamination in cell passage processes.

Description

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Claims

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Application Information

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Owner CHINA JILIANG UNIV
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