Short Notes on cryopreservation|Biotechnify|BTF

 Short notes on cryopreservation|Biotechnify|BTF

What is cryopreservation?

Cryo-preservation or cryo-conservation is a process of cooling and storing cells, tissues, or organs at very low temperatures to maintain their viability. For example, the technology of cooling and storing cells at a temperature below the freezing point ('196' C) permits high rates of survivability of the cells uponthawing. where organelles, cells, tissues, extracellular matrix, organs, or any other biological constructs susceptible to damage caused by unregulated chemical kinetics are preserved by cooling to very low temperatures^[1] (typically −80 °C using solid carbon dioxide or −196 °C using liquid nitrogen). At low enough temperatures, any enzymatic or chemical activity which might cause damage to the biological material in question is effectively stopped.

Explain :-

 Cryopreservation technique is used to freeze and thaw the eggs that can be used in IVF. Highest Success Rate. Cost Effective Fertility. Services: IUI, ICSI, IVF, Infertility Workup, Laser Assisted Hatching, Cryopreservation.

Cryopreservation is the method of keeping the live cells, tissues and other biological samples in a deep freeze at subzero temperatures for the storage or preservation. The sample is commonly kept at −196°C.

At such low temperatures, all the biological activities of the cells stop and the cell dies. Cryopreservation helps the cells to survive freezing and thawing.

Cryopreservation is also used to freeze and store human embryos and sperm. It is especially valuable for the freezing of extra embryos that are generated by in vitro fertilization (IVF).

Cryopreservation Process/procedure

In this process, biological materials including cells, oocytes, spermatozoa, tissues, ovarian tissues, pre-implantation embryos, organs, etc. are kept in extremely cold temperatures without affecting the cell’s viability.

Dry Ice and liquid nitrogen are generally used in this method.

Cryopreservation is the use of very low temperatures to preserve structurally intact living cells and tissues for a long period of time.

Depending on the cell types or given cells among different mammalian species, there is great diversity in cryobiological response and cryosurvival during the freezing and thawing cycle .

Cryopreservation processes can generally be grouped into the following types: (1)slow freezing; (2) vitrification, which involves the solidification of the aqueous milieu of the cell or tissue into a noncrystalline glassy phase; (3) subzero nonfreezing storage; and (4) preservation in the dry state.

Generally, the storage of mammalian cells in the dry state is not readily possible because of difficulties in introducing the disaccharide trehalose (disaccharide of glucose, 342 Da)and amino acids (used as preservatives in plants) into the intracellular region.

The major steps in cryopreservation are (1): the mixing of CPAs with cells or tissues before cooling; (2) cooling of the cells or tissues to a low temperature and its storage; warming of the cells or tissues; and removal of CPAs from the cells or tissues after thawing.

The appropriate use of CPAs is therefore important to improve the viability of the sample to be cryopreserved.

Cryopreservation is based on the ability of certain small molecules to enter cells and prevent dehydration and formation of intracellular ice crystals, which can cause cell death and destruction of cell organelles during the freezing process. 

Concept :

Cryopreservation of Sperm

The semen sample is mixed with a solution, which provides protection during freezing and thawing. Followed by transfer to plastic vials, which are then kept in liquid nitrogen for freezing.

Cryopreservation of Embryos

During the infertility treatment, hormones are used to stimulate the development of eggs. The eggs are then taken out and fertilized in the lab. More embryos can be created and transplanted to the woman’s uterus. These embryos can be cryopreserved and can be used at some later date. By this, the female can get an additional transfer of embryo in future, without spending on another IVF cycle.

What is the temperature of cryopreservation?

Cryopreservation may be defined as the maintenance of biologics at sub-freezing temperatures, below −80°C and typically below −140°C.

Why is liquid nitrogen used in cryopreservation?

The use of liquid nitrogen is an effective long-term method for storing viable samples while maximizing energy efficiency and providing an environmentally friendly approach to cryopreservation. This innovative freezing method ensures that cells remain viable, and indefinite storage is possible.

Applications of Cryopreservation

Cryopreservation or basically we can say conservation meghod is a long-term storage technique, which is mainly used for preserving and maintaining viability of the biological samples for a longer duration and This preservation technique is widely used in cryosurgery, molecular biology, ecology, food science, plant physiology, and in many medical application.

 Some other applications :

1. Gene Bank.

2. Blood transfusion.

3. In vitro fertilization.

4. Artificial insemination.

5. Storage of rare germplasm.

6. Seed Bank

Etc.,..

What are the advantages of cryopreservation?

Cryopreservation is a long-term storage technique with very low temperatures to preserve the structurally intact living cells and tissues for extended period of time at a relatively low cost. Cryopreservation is to preserve and store the viable biological samples in a frozen state over extended periods of time.

_____________________________________________

Some interesting things  🤠

:- if you are searching top ranking University in india then click next and see or many colleges..

Short notes on X-ray Crystallography | BTF

 X-ray crystallography is the most powerful method to obtain a macromolecular structure.

X-ray crystallography is a tool used for determining the atomic and molecular structure of a crystal. The underlying principle is that the crystalline atoms cause a beam of X-rays to diffract into many specific directions. By measuring the angles and intensities of these diffracted beams, a crystallographer can produce a 3D picture of the density of electrons within the crystal.

X-Ray crystallography is a tool used to provide structural information about molecules. The technique was developed in 1912 by William Henry Bragg and William Lawrence Bragg (a father and son team who won the 1915 Nobel Prize in Physics for their work in the field), who built upon earlier work by Max von Laue.

X-ray crystallography remains the most robust method to determine protein structure at the atomic level. However, the bottlenecks of protein expression and purification often discourage further study. 

X-ray crystallography  is the experimental science determining the atomic and molecular structure of a crystal, in which the crystalline structure causes a beam of incident X-rays to diffract into many specific directions. By measuring the angles and intensities of these diffracted beams, a crystallographer can produce a three-dimensional picture of the density of electrons within the crystal. From this electron density, the mean positions of the atoms in the crystal can be determined, as well as their chemical bonds, their crystallographic disorder, and various other information.

What is X ray diffraction simple definition?

: a scattering of X-rays by the atoms of a crystal that produces an interference effect so that the diffraction pattern gives information on the structure of the crystal or the identity of a crystalline substance.

X-ray crystallography is a technique used for determining the high-resolution, three-dimensional crystal structures of atom and molecules and has been fundamental in the development of many scientific fields. In its first decades of application, it is mainly used for determining the size of atoms, the lengths and types of chemical bonds, the atomic-scale differences among various materials, as well as the crystalline integrity, grain orientation, grain size, film thickness and interface roughness of the related materials, especially minerals and alloys.

 

Through X-ray crystallography, the chemical 

structure of thousands of organic, inorganic, 

organometallic, and biological compounds 

are determined every year. 

Bragg 

Spectrometer :

X-ray was the name given to the 

highly penetrating rays which 

emanated when high energy 

electrons struck a metal target. 

What is the principle of X ray crystallography?

X-ray crystallography is a tool used for determining the atomic and molecular structure of a crystal. The underlying principle is that the crystalline atoms cause a beam of X-rays to diffract into many specific direction.  

What is X ray crystallography used for?

X-ray crystallography is a tool used for determining the atomic and molecular structure of a crystal. The underlying principle is that the crystalline atoms cause a beam of X-rays to diffract into many specific direction. 

Why is Xray crystallography important?

X-ray crystallography enables the identification of the atomic and molecular structure of a crystal. ... The X-ray technique provides direct structural information on molecules at the atomic level and is recognized as a reliable structure determination method (Gaudencio and Pereira, 2015).

Where is Xray crystallography used?

Most scientists use x-ray Crystallography to solve the structures of protein and to determine functions of residues, interactions with substrates, and interactions with other proteins or nucleic acids. Proteins can be co - crystallized with these substrates, or they may be soaked into the crystal after crystallization. 

X-ray crystallography is also a routine technique to determine how a drug interacts with its target and what modifications could improve the interaction. Prakasham et al.76 have investigated diastase enzyme immobilized on nickel-impregnated silica paramagnetic NPs and they characterized them by FTIR and X-ray crystallography.

Why do we study crystallography?

Crystallography is the study of atomic and molecular structure. Crystallographers want to know how the atoms in a material are arranged in order to understand the relationship between atomic structure and properties of these materials. Crystallography began with the study of crystals, like quartz. 

 

Applications of X-ray crystallography

X-ray crystallography is used to analyze many different molecules and has been used in many famous projects in the fields of organic and inorganic chemistry. Early structures which were resolved using the technique were simple crystals, including quartz. As well as the analysis of organic molecules (proteins, vitamins, nucleic acids) and inorganic molecules and structures, X-ray crystallography has been used to develop novel materials in both material and life sciences.

Drug identification

Investigation of bones

Characterization of textile fibers and polymers

Integrated circuits

Short notes on X-ray Crystallography | BTF

Fermentation |Biotechnify |BTF

 

Fermentation : Definition & Introduction 

The chemical breakdown of a substance by bacteria, yeasts, or other microorganisms, typically involving effervescence and the giving off of heat.

"the fermentation of organic matter by microorganisms in the gut"

Fermentation is a metabolic process that produces chemical changes in organic substrates through the action of enzymes. In biochemistry, it is narrowly defined as the extraction of energy from carbohydrates in the absence of oxygen.

The science of fermentation is known as zymology.

In microorganisms, fermentation is the primary means of producing adenosine triphosphate (ATP) by the degradation of organic nutrients anaerobically.[2] Humans have used fermentation to produce foodstuffs and beverages since the Neolithic age. For example, fermentation is used for preservation in a process that produces lactic acid found in such sour foods as pickled cucumbers, kombucha, kimchi, and yogurt, as well as for producing alcoholic beverages such as wine and beer. Fermentation also occurs within the gastrointestinal tracts of all animals, including humans.

The word "ferment" is derived from the Latin verb fervere, which means to boil.it is all about in etymology.

Fermentation, chemical process by which molecules such as glucose are broken down anaerobically. More broadly, fermentation is the foaming that occurs during the manufacture of wine and beer, a process at least 10,000 years old. The frothing results from the evolution of carbon dioxide gas, though this was not recognized until the 17th century.

How Does Fermentation Work?

Microorganisms survive using carbohydrates (sugars, such as glucose) for energy and fuel.

Organic chemicals like adenosine triphosphate (ATP) deliver that energy to every part of a cell when needed.

Microbes generate ATP using respiration. Aerobic respiration, which requires oxygen, is the most efficient way to do that. Aerobic respiration begins with glycolysis, where glucose is converted into pyruvic acid. When there’s enough oxygen present, aerobic respiration occurs.

Fermentation is similar to anaerobic respiration—the kind that takes place when there isn’t enough oxygen present. However, fermentation leads to the production of different organic molecules like lactic acid, which also leads to ATP, unlike respiration, which uses pyruvic acid.

Depending upon environmental conditions, individual cells and microbes have the ability to switch between the two different modes of energy production.

Organisms commonly obtain energy anaerobically through fermentation, but some systems use sulfate as the final electron acceptor in the electron transport chain.

Fermentation is all down to the actions of tiny natural microbes, who colonize and cultivate everything from our digestive systems, to this colorful spring in Yellowstone seen in the picture above, to the food and drink we eat. 

Microbes use carbohydrates (sugars, such as glucose) for energy to fuel their survival. To make use of that energy, organic chemicals like adenosine triphosphate (ATP) deliver it when needed to every part of a cell.

Microbes - and our own body cells - use respiration to generate ATP. The most efficient way for them to do that is through a process known as aerobic respiration, which requires oxygen.

Aerobic respiration starts with glycolysis, where glucose is converted into pyruvic acid. Then, when there's enough oxygen around, aerobic respiration takes place. 

Fermentation is similar to the kind of respiration that takes place when there isn't enough oxygen present, namely anaerobic respiration. However unlike respiration, which uses pyruvic acid, fermentation leads to the production of different organic molecules like lactic acid, which also leads to ATP.  

What are three types of fermentation? 

Lactic acid fermentation

Lactic acid fermentation is a metabolic process by which glucose and other six-carbon sugars (also, disaccharides of six-carbon sugars, e.g. sucrose or lactose) are converted into cellular energy and the metabolite lactate, which is lactic acid in solution.

Yeast strains and bacteria convert starches or sugars into lactic acid, requiring no heat in preparation. These anaerobic chemical reactions, pyruvic acid uses nicotinamide adenine dinucleotide + hydrogen (NADH) to form lactic acid and NAD+. (Lactic acid fermentation also occurs in human muscle cells. During strenuous activity, muscles can expend adenosine triphosphate (ATP) faster than oxygen can be supplied to muscle cells, resulting in lactic acid buildup and sore muscles. In this scenario, glycolysis, which breaks down a glucose molecule into two pyruvate molecules and doesn’t use oxygen, produces ATP.) Lactic acid bacteria are vital to producing and preserving inexpensive, wholesome foods, which is especially important in feeding impoverished populations. This method makes sauerkraut, pickles, kimchi, yogurt, and sourdough bread.

Ethanol fermentation/alcohol fermentation

Yeasts break pyruvate molecules—the output of the metabolism of glucose (C6H12O6) known as glycolysis—in starches or sugars down into alcohol and carbon dioxide molecules. Alcoholic fermentation produces wine and beer.

Ethanol fermentation, also called alcoholic fermentation, is a biological process which converts sugars such as glucose, fructose, and sucrose into cellular energy, producing ethanol and carbon dioxide as by-products.

Acetic acid fermentation

Starches and sugars from grains and fruit ferment into sour tasting vinegar and condiments. Examples include apple cider vinegar, wine vinegar, and kombucha

Cell Culture Techniques |Biotechnify|BTF

What is Cell Culture? 

Cell culture is the process by which cells are grown under controlled conditions, generally outside their natural environment. After the cells of interest have been isolated from living tissue, they can subsequently be maintained under carefully controlled conditions.

Essentially , cell culture involves the distribution of cells in an artificial environment (in vitro) which is composed of the necessary nutrients, ideal temperature, gases, pH and humidity to allow the cells to grow and proliferate.

• In vivo - When the study involves living biological entities within the organism.

• In vitro - When the study is conducted using biological entities (cells, tissue etc) that has been isolated from their natural biological environment. E.g. tissue or cells isolated from the liver or kidney. 

Cell culture refers to the removal of cells from an animal or plant and their subsequent growth in a favorable artificial environment. The cells may be removed from the tissue directly and disaggregated by enzymatic or mechanical means before cultivation, or they may be derived from a cell line or cell strain that has already been established.

What is Cell Culture Techniques ?

In cell culture techniques, cells (or tissues) are removed from a plant or an animal and introduced into a new, artificial environment that can support their proliferation (survival and growth). Some of the requirements of such an environment for the proliferation of the cells include: A substrate (source of nutrition).

Cell culture techniques play a key role in the development of new anticancer drugs by imposing additional constraints on those of receptor interaction alone, such as drug uptake and efflux, interaction with other cellular receptors, and cellular metabolism

Primary culture :

Primary culture refers to the stage of the culture after the cells are isolated from the tissue and proliferated under the appropriate conditions until they occupy all of the available substrate (i.e., reach confluence). At this stage, the cells have to be subcultured (i.e., passaged) by transferring them to a new vessel with fresh growth medium to provide more room for continued growth

Cell Culture Protocol :

Cell culture protocols are meant to ensure that culture procedures are carried out to the required standards. This is not only meant to prevent the contamination of the cells, but to also ensure that the researchers themselves are protected from any form of contamination.

However, the nature of the work is expected to conform to the appropriate ethical guidelines. Therefore, before anything else, it is essential to ensure that the entire procedure conforms with both medical-ethical and animal- experiment guidelines. This is because going against such legislation and guidelines can result in heavy penalties and even shutting down of the laboratory.

Before start , carry out the following procedure:

Ensure that the working are is sanitized (using 70 percent ethanol)Always use a new pair of gloves. If a pair of gloves has to be used for another cell culture procedure, they should be sanitized using 70 percent ethanol and allowed to air dry.

Any equipment that had been taken out of the cabinet should also be sanitized to prevent any contamination such equipment as pipette, glass jars and plastics to be used for the procedure should be autoclaved

Although there are a wide range of culture media for cells, it is important to keep in mind that cell cultures, and particularly primary cell cultures are easily prone to contamination in addition to the risk of containing undetected viruses. For this reason, all material should be handled as potentially infectious in order to avoid any infections.

Protocols for cell culture preparation :

Always check the information on the container to ensure that the medium is appropriate for the cell to be cultured,

Once prepared, the cell culture should be maintained under the recommended temperature range,

Monitor the culture every 30- 48 hours and check for confluency (when cells completely cover the surface of the culture) - However, this is largely dependent on the type of cells.

Once the procedure is completed and the cells have been analyzed, the culture should be appropriately discarded. Here, it is important to take a lot of caution given that by this time, cells have already proliferated and increased in numbers. Moreover, there are high chances that the specimen has been contaminated, which increase the risks of causing infections to the researcher if not handled appropriately.

What is Cell Culture used for? 

Cell culture is one of the major tools used in cellular and molecular biology, providing excellent model systems for studying the normal physiology and biochemistry of cells (e.g., metabolic studies, aging), the effects of drugs and toxic compounds on the cells, and mutagenesis and carcinogenesis.

What is Cell Passaging? 

Subculturing, also referred to as passaging cells, is the removal of the medium and transfer of cells from a previous culture into fresh growth medium, a procedure that enables the further propagation of the cell line or cell strain.

Applications of Cell Culture :

Cell culture is one of the major tools used in cellular and molecular biology, providing excellent model systems for studying the normal physiology and biochemistry of cells (e.g., metabolic studies, aging), the effects of drugs and toxic compounds on the cells, and mutagenesis and carcinogenesis. It is also used in drug screening and development, and large scale manufacturing of biological compounds (e.g., vaccines, therapeutic proteins). The major advantage of using cell culture for any of these applications is the consistency and reproducibility of results that can be obtained from using a batch of clonal cells.

Conclusion :

In conclusion, cell culture is an indispensable tool in modern day medicine and its applications are innumerable in diagnosis of human infection. Cell culture methods are unbiased to some extent and only limited by the ability of the virus to grow in a particular cell line.

Isolation Techniques - short explanation | notes on isolation techniques

 Isolation Techniques - short explanation | notes on isolation techniques

Isolation Techniques 

1. the process of separating, or the state of being alone.

2. the physiologic separation of a part, as by tissue culture or by interposition of inert material.

3. the extraction and purification of a chemical substance of unknown structure from a natural source.

4. the separation of infected individuals from those uninfected for the period of communicability of a particular disease; see also quarantine.

5. the separation of an individual with a radioactive implant from others to prevent unnecessary exposure to radioactivity.

Definition :

Isolation refers to the precautions that are taken in the hospital to prevent the spread of an infectious agent from an infected or colonized patient to susceptible persons.

In microbiology, the term isolation refers to the separation of a strain from a natural, mixed population of living microbes, as present in the environment, for example in water or soil flora, or from living beings with skin flora, oral flora or gut flora, in order to identify the microbe(s) of interest.

History :

The laboratory techniques of isolating microbes first developed during the 19th century in the field of bacteriology and parasitology using light microscopy. Proper isolation techniques of virology did not exist prior to the 20th century. The methods of microbial isolation have drastically changed over the past 50 years, from a labor perspective with increasing mechanization, and in regard to the technologies involved, and with it speed and accuracy.

Purpose

Isolation practices are designed to minimize the transmission of infection in the hospital, using current understanding of the way infections can transmit. Isolation should be done in a user friendly, well-accepted, inexpensive way that interferes as little as possible with patient care, minimizes patient discomfort, and avoids unnecessary use.

Precautions Types

There are three types of transmission-based precautions--contact, droplet, and airborne - the type used depends on the mode of transmission of a specific disease.

Precautions :

The type of precautions used should be viewed as a flexible scale that may range from the least to the most demanding methods of prevention. These methods should always take into account that differences exist in the way that diseases are spread. Recognition and understanding of these differences will avoid use of insufficient or unnecessary interventions.

Standard precautions :

Standard Precautions define all the steps that should be taken to prevent spread of infection from person to person when there is an anticipated contact with:

Blood

Body fluids

Secretions, such as phlegm

Excretions, such as urine and feces (not including sweat) whether or not they contain visible blood

Nonintact skin, such as an open wound

Mucous membranes, such as the mouth cavity.

Standard Precautions includes the use of one or combinations of the following practices. The level of use will always depend on the anticipated contact with the patient:

Handwashing, the most important infection control method

Use of latex or other protective gloves

Masks, eye protection and/or face shield

Gowns

Proper handling of soiled patient care equipment

Proper environmental cleaning

Minimal handling of soiled linen

Proper disposal of needles and other sharp equipment such as scalpels

Placement in a private room for patients who cannot maintain appropriate cleanliness or contain body fluids.

Transmission based precautions :

Transmission Based Precautions may be needed in addition to Standard Precautions for selected patients who are known or suspected to harbor certain infections. These precautions are divided into three categories that reflect the differences in the way infections are transmitted. Some diseases may require more than one isolation category.

AIRBORNE PRECAUTIONS. Airborne Precautions prevent diseases that are transmitted by minute particles called droplet nuclei or contaminated dust particles. These particles, because of their size, can remain suspended in the air for long periods of time; even after the infected person has left the room. Some examples of diseases requiring these precautions are tuberculosis, measles, and chickenpox.

Following isolation methods are employed to isolate microbes from mixed cultures:

1. Streaking

2. Plating

3. Dilution

4. Enriched procedure, and

5. Single cell technique.

Description :

Isolation practices can include placement in a private room or with a select roommate, the use of protective barriers such as masks, gowns and gloves, a special emphasis on handwashing (which is always very important), and special handling of contaminated articles. Because of the differences among infectious diseases, more than one of these precautions may be necessary to prevent spread of some diseases but may not be necessary for others.The Centers for Disease Control and Prevention (CDC) and the Hospital Infection Control Practice Advisory Committee (HICPAC) have led the way in defining the guidelines for hospital-based infection precautions. The most current system recommended for use in hospitals consists of two levels of precautions. The first level is Standard Precautions which apply to all patients at all times because signs and symptoms of infection are not always obvious and therefore may unknowingly pose a risk for a susceptible person. The second level is known as Transmission-Based Precautions which are intended for individuals who have a known or suspected infection with certain organisms.

Frequently, patients are admitted to the hospital without a definite diagnosis, but with clues to suggest an infection. These patients should be isolated with the appropriate precautions until a definite diagnosis is made.

Notes :

It is absolutely essential that you sterilize your loop between each streaking, either by using the incinerator or by obtaining a new sterile plastic loop. This is the most common mistake students make.

Don’t leave your plate open too long or extra bacteria from the environment will fall into your plate.

Do not be disappointed if you do not get isolated colonies on your first try. This is a difficult procedure.