GENETIC TEACHER

Steps uses of Light Microscope: 1- Turn On light source, 2- Turn the objective lens to low, 3- Lower the objective lens without touch the slide, 4- Center the image by moving the stage, 5- Adjust the diaphragm for optimum light, 6- Adjust focus first with coarse then fine knob #geneticteacher

For Staining Eukaryotic Cells: 1- Use toothpick to gently scarp the inner lining of check, 2- Put the scarping on the clean glass slide and add one drop of methylene blue, 3- Cover with cover slip and observe under the light microscope #geneticteacher

Precaution During Work with Light Microscope: 1- Ensure working area clean and tidy, 2- Wear gloves when handling immersion oil and stains, 3- Clean lens gently with lens tissue immediately after use, 4- After finish leave the low power objective lens in place and lower the stage, 5- Turn off light source, 6- All glass slides must be disposed into sharps container #geneticteacher

Structure Functions of Light Microscope: 1- Eyepiece or Ocular Lens (Located on the top of the body tube to magnifies the specimen 10X). 2- Objective Lenses (Located on the nose piece at the bottom of the body tube and used in combination with the eyepiece to provide a range of magnification, so, objective lenses 8X or 10X (Shorter Lens) and 100X (Longest Lens) in normal, therefore, (8X= Obtain initial focus on a sample to scan a slide for a point of interest), (10X= Examine any finding closely like mitosis and meiosis divisions), (40X= Examine any finding in a greater details), (100X= Examine in more details the findings), so, for scanning and low power use coarse and fine knob and for high power use fine knob only. 3- Nose-Piece (Located at the bottom of the body tube to hold objective lenses and rotates to enable magnification). 4- Arm and Base (Carry and Support the upper parts and whole body of the microscope by adding one hand on the arm and the other hand on the base). 4- Stage...

Light Microscope use to visualize the internal organs of eukaryotic cells bey enlarging images up to 1500 time their original size to identify bacterial cells with distinctive shapes and to detect certain microbial elements such as bacterial endospores, so, Micro=Small and Scope=Look at #geneticteacher

For Handling Light Microscope: 1- Carry light microscope properly, 2- Always begin focusing with 4X or 8X objective lens by using coarse, 3- Use Coarse Knob to focus and then fine adjustment knob to make the image crystal clear, 4- Use one drop immersion oil only with 100X objective lens, 5- After finishing lower the stage and remove the slide then clean the microscope with lens paper tissue, but be sure the stage is at its lowest position before putting the microscope away, 6- Turn off the light before putting the microscope away, 7- wrap the cord correctly before putting the microscope away, 8- Return the microscope to the correct cabinet #geneticteacher

HPLC Applications: 1- Chemical Sciences (Polystyrenes and Dyes and Phthalates), 2- Biosciences (Proteins and Peptides and Nucleotides), 3- Consumer Products (Lipids and antioxidants and Sugars), 4- Environmental Sciences (Polyaromatic Hydrocarbon and Inorganic Ions and Herbicides), 5- Pharmaceutical (Tetracycline and Corticosteroid and Antidepressants and Barbiturates), 6- Clinical (amino Acids and Vitamins and Homocysteine) #geneticteacher

Chromatogram is a graph show the detector response as a function of elution time in HPLC device. Retention Time is a time for each component after injection of the mixture until component reaches the detector of HPLC #geneticteacher

Mechanisms of HPLC: 1- Sample is introduced through column, 2- Sample components moves to column with high pressure delivered from pump for separating, 3- Separated components detects by detector to measure their accounts in chromatogram. Therefore, 1- Sample size must be tiny (3-5) micrometer in diameter, 2- Internal diameter of column must be small with high sensitivity, 3- Pump pressure must be high for high separation, 4- Temperature must be high for high separation #geneticteacher

HPLC or High Speed Liquid Chromatography or High Pressure Liquid Chromatography or High Performance Liquid Chromatography used to separate or identify or quantify each component in mixture in stationary and mobile phases through column Chromatography with high pressure, so, the heart of HPLC system is column. Hence, Revisor holds solvent, Pump generate flow, Injector insert solvent with sample into stream in high pressure through column, Detector identify an quantify separated components in chromatogram frame #geneticteacher

Types of Chromatography : 1- Column Chromatography (Sample held directly through top column under gravity effects), 2- Paper Chromatography (Separate substances by cellulose paper as chromatographic sorbent), 3- Thin Layer Chromatography (Sheet of glass or metal or plastic coated with thin layer of solid adsorbent such as silica or alumina), 4- Gas Chromatography (Describe analytical separation in gas phase) #geneticteacher

Chromatography is a technique that separate the components of solutes of a mixture on the basis of the relative amounts of each solute distributed between fluid liquid or gas stream (Mobile Phase) and contiguous liquid or solid stream (Stationary Phase). #geneticteacher

There are five steps of pH Meter Calibration: 1- Place electrode of pH Meter in the (buffer solution 7) then read, 2- Rinse electrode with distilled water, 3- Place electrode in the (buffer solution 4) then read, 4- Rinse electrode in distilled water, 5- Clean electrode with tissue after use #geneticteacher

Types of ELISA-Plate on the basis of binding structure between antibody and antigen: 1- Direct ELISA (Involve attachment of antigen to solid phase follow by enzyme labeled antibody), 2- Indirect ELISA (Involve attachment of antigen to solid phase but the primary antibody isn’t labeled and secondary antibody is labeled with enzyme), 3- Sandwich ELISA (Involve attachment of capture antibody to solid phase follow by enzyme labeled antibody), 4- Competitive ELISA (Involve simultaneous addition of competing antibodies that decrease signal of samples when add secondary antibody) #geneticteacher

Steps Procedures of ELISA-PLATE: 1- Add Antigen, 2- Wash with PBST, 3- Add Primary Antibody, 4- Wash with PBST, 5- Add Secondary Antibody, 6- Wash with PBST, 7- Add Substrate for Enzyme, 8- Observe color development #geneticteacher

ELISA-Plate is a flat bottom polystyrene plate contains 96 or 384 wells holding 350 microliters each. Immunoassay is a laboratory technique use to bind between antigen and its homologous antibody to identify specific antigen or antibody in the sample. Antibodies are proteins produced in response to antigenic simulations. Antigen mean any molecule that elicit the production of antibodies when introduced into body. Adsorption is a process of adding antigen or/and antibody and samples with dilution buffer to soldi phase through incubation. Enzyme Conjugate mean enzyme that attached irreversibly to antibody. Washing mean flooding ELISA wells with Phosphate Buffer Saline Tween (PBST) three times to separate bound from unbound reagents. ELISA Plate used for (Microbiology Test – Food Allergens Test – Diseases Test) #geneticteacher

Enzyme Linked Immunosorbent Assay (ELISA-Plate) have polystyrene plate (96 or 384 wells) and require microliter quantities, so, it widely used in immunoassay by immobilize antigen to sild surface and linked complexes antigen/antibody to an enzyme. Therefore, detection is accomplished by assessing the conjugated enzyme activity by incubation with substrate to produce measurable product at 450 nanometers. So, during use ELISA-Plate use 50 microliters positive control (+) in first three wells and use 50 microliters negative control (-) in second three wells and use 50 microliters each samples in duplicate or triplet in remaining wells #geneticteacher

For pH Meter Calibration use distilled water to rinse electrode, then, place the electrode in pH=7 buffer solution and gently dob dry using non-abrasive tissue. After that, wait about one minute for pH Meter Stabilization, then, adjust the pH Meter until read 7. Next, remove the electrode from the buffer solution 7, then, clean it by rinsing with distilled and gently dob dry using non-abrasive tissue. After that, place the electrode in a buffer solution 4, then, wait about one minute for Stabilization, then, adjust the pH Meter until read 4. After that, remove the electrode from buffer solution 4, then, clean it by rinsing with distilled water and gently dob dry using non-abrasive tissue. Finally, Rinse and save pH Meter electrode with distilled water inside beaker without hitting beaker bottom #geneticteacher

The standard buffer that are used in pH Meter Calibration are solution that have constant pH values used to calibrate pH Meter from electrodes. The most commonly used of standard buffer have pH (4 and 7 and 10). Hence, If your plane to measure pH in acidic solutions use pH=4 buffer, If your plane to measure pH in high solutions use pH=10 buffer, If you want to be able to measure pH in wider range use all buffers #geneticteacher

pH Meter is a scientific instrument that measures hydrogen ions activity in water based solutions indicating its acidity of alkalinity expressed as pH. So, pH is a unit of measure which describes the degree of acidity or alkalinity of solution. Therefore, the scale of pH Meter range from (0 – 14): 1- If the pH less than 7 the solution will be acid, 2- If the pH greater than 7 the solution will be basic, 3- If the pH equal 7 the solution will be Neutral #geneticteacher

Radiation Sterilization: 1- Ionizing Radiation such as X-rays and Gamma rays that sterilize syringes and gloves and plastic tubes and dishes, 2- Non-ionizing Radiation such as Infrared and Ultraviolet that sterilize bench work of laminar air flow cabinet #geneticteacher

Physical Sterilization and Chemical Disinfection that are kill most forms of microorganisms by control growth and metabolisms. Physical sterilization consist of: 1- Heat sterilization (Dry Heat + Moist Heat), 2- Radiation Sterilization (Non-ionizing Radiation + ionizing Radiation), 3- Filter Sterilization (Depth Filter + Membrane Filter + Air Filter). Chemical Disinfection (70% Alcohol + 70% Dettol + 2% Iodine (Betadine) + 5% Chlorine Compounds + 37% Formaldehyde (Formalin) + 5% Phenol Group + 3% Oxidizing Agent + 2% Ethylene Oxides + 2% Glutaraldehyde). Sterile Body Site Called Sterile. Non-Sterile Body Site Called Disinfected or Antiseptic #geneticteacher

Moist Heat Sterilization: 1- Sterilization Below 100 degree Celsius such as Pasteurization that sterilize milk and juice to inactive harmful microorganisms , 2- Sterilization at 100 degree Celsius such as Water Bath sterilization that sterilize serum and vaccine at 60 degree Celsius for one hour several days and Boiling that sterilize physical objects in a boiling water at 100 degree Celsius for 10 to 30 minutes and Tyndalization or steaming by using boiling water vapor for two minutes three days sequences , 3- Sterilization above 100 degree Celsius such as Autoclave that sterilize physical items for 20 minutes to coagulate proteins and destroy functional integrity of cell membrane of microorganisms #geneticteacher

Filtration Sterilization: (1- Depth Filter, 2- Membrane Filter, 3- Air Filter) to trap and reduce microorganisms cells from liquid solutions using 0.22 micrometer for bacteria or 10 millimeters for viruses #geneticteacher

Dry Heat Sterilization: 1- Red Heat Sterilization (Sterilize inoculation loops and needles and forceps and scissors in redness hot over Bunsen flam) , 2- Flaming Heat sterilization (Sterilize mouth of culture tubes and flasks and glass slides over Bunsen flam as well without redness hot), 3- Incineration Heat Sterilization (Destroy pathological materials until become ash using fire with forced air blower), 4- Hot Air Over Sterilization (sterilize glassware and instruments and powder without water at 160 at 180 degree Celsius in about two hours to destroy oxidative processes of microorganism) #geneticteacher

Stable Incubator have tow doors made of glass to easily view contents for good regulations of temperatures and humidity for fast recovery times after open and close door #geneticteacher

Shaking Incubator have combined shaker and incubator to provide rapid and uniform artificial conditions for aeration cultures such as cell culture and cell aeration and solubility studies by evenly mix nutrients cultures #geneticteacher

Incubator Precautions: 1- Don’t touch plug tight into socket with wet hand, 2- Don’t put anything heavy on the power code to avoid electrical shock, 3- Don’t place any explosion or flammable materials beside or inside incubator, 4- Don’t expose incubator to direct sun rise to avoid damage, 5- Don’t put water above incubator body to avoid electric shock, 6- Don’t set incubator in part of wet materials, 7- Don’t leaves door open during normal operation to avoid contamination, 8- Don’t place incubator near autoclaves or hot air ovens to avoid too much heat, 9- Ensure proper spacing between samples and chamber walls to avoid longer time incubation, 10- Place incubator on top of dry and stable surface, 11- Place incubator in good temperature room and humidity for best performance, 12- Use soft clothes for cleaning without solvents, 13- Ensure correct connect power electricity to prevent electrical hazard, 14- Make circular maintenance every year for good performance #geneticteacher

Types of Incubators: 1- Stable Incubator (Provide ideal culture conditions with high resistant contamination), 2- Shaking Incubator (Used for aeration cultures such as cell culture and cell aeration and solubility studies) #geneticteacher

Microbiological Incubator is an insulate heating and cooling cabinet use to grow and maintain microbiological cultures under artificial conditions at constant and desired temperature by maintain optimal temperature and humidity and carbon dioxide and oxygen contents inside chamber. So, good temperature for most bacteria (35 – 37) degree Celsius and good temperature for most fungi 25 degree Celsius #geneticteacher

For cleaning Incubator: 1- Remove Shelves, 2- Remove dirt by using mild soap solution and sponge, 3- Rinse soap solution twice with distilled water, 4- Dry all rinsed parts with paper towel, 5- Wipe surfaces with 70% ethanol. In addition, before working with incubator set required parameters and place sterile water under death stable incubator shelves to prevent culture media from drying out and avoid open cabinet door repeatedly #geneticteacher

Incubator must be place for (18 – 25) degree Celsius on flat and stable surface in at least 1.5 meter from light and 20 cm from walls #geneticteacher