Introduction to Animal Science Lab Animal 1 PDF

HELWAN NATIONAL UNIVERSITY FACULTY OF SCIENCE BIOTECHNOLOGY & GENETIC ENGINEERING PROGRAM Introduction to animal science First year students 2024-2025  MICROSCOPY What is microscopy ? Technical field using...

HELWAN NATIONAL UNIVERSITY FACULTY OF SCIENCE BIOTECHNOLOGY & GENETIC ENGINEERING PROGRAM Introduction to animal science First year students 2024-2025  MICROSCOPY What is microscopy ? Technical field using microscopes to view samples and objects that can’t be seen with unaided eyes. Micro = Microscope is an small instrument used to study Microscope very small organisms or Scope = particles which aren’t application visible by naked eyes.  Why we use microscopes ? Human naked eye can see smallest object of 0.1 mm only, but microscopes can see much smaller objects. Anton Van Leeuwenhoeck Anton Van Leeuwenhoeck made the microscope for first time in 1674.  Types of microscopes Light microscope (LM) Electron microscope (EM) (uses light as a source of images) (uses electron beam as a source of images) Scanning electron microscope Simple LM Compound LM (SEM) Gives a 3D images for samples. Image shows the external morphology of sample. Transmission electron Simple Monocular Biocular microscope (TEM) light compound compound Gives a 2D images for samples. microscope LM has 2 LM has 2 Image shows the internal structure has only sets of lens sets of lens of sample. one lens with one with two eye piece eye pieces  Light microscope Monocular compound light Biocular compound light Simple light microscope microscope microscope The most common used microscopes  The parts of compound Light microscope Optical Mechanical system system Eye piece tube Ocular lens (eye piece) Ocular lens (eye piece) Head Objective lens Nose piece Arm Mechanical stage Mechanical stage Aperture Coarse adjustment Fine adjustment Diaphragm Stage controls Base Condenser Illuminator (light) Light switch Brightness adjustment  Lenses of compound light microscope Scanning Low power High power Oil immersion lens lens lens lens Using of this lens by using a special Most common oil to show the used lenses specimen Sliding ruler X and Y axis It is used to adjust and save the location of slide on stage of microscope  Electron microscope Scanning electron microscope Transmission electron (SEM) microscope (TEM) It creates a 3D image by detecting reflected It creates a 2D image using electrons that are electrons passing through the sample  Comparison between scanning and transmission electron microscopes Features Scanning electron microscope Transmission electron microscope (SEM) (TEM) Source of Electrons are used for image Electrons are used for image illumination formation formation Types of visualized Specimens are coated with gold, the Thin section of specimen are used, cells electrons are reflected back gives the electrons pass through section image for surface of specimen gives image for internal structure of specimen Images 3D images 2D images Nature of lenses one electrostatic lens with few one electrostatic lens with few electromagnetic lenses electromagnetic lenses Medium Vacuum Vacuum  Concepts Resolution power It is the ability of a microscope to distinguish the details of the sample It is the ability of a lens to distinguish small objects that are close together Magnification power The enlargement of images up to 1000 times of its original size Working distance The vertical distance between the front surface of lens and surface of cover glass (on specimen) Numerical aperture (NA) The ability of an objective lens to collect light NA= n sinθ Angle of light Refractive index collected by the Numerical of medium (n=1 objective lens aperture in air, n=1.5 in oil) ‫الرقم اللي فوق على‬ ‫اليمين‬  Wave length Magnification (λ = 0.527) power of eye piece (ocular lens) λ.0.6 R= NA Resolution power Numerical aperture Power of eye piece (ocular lens) Power of Magnification power of objective lens Magnification objective lens )‫(الرقم اللي فوق على الشمال‬ M= (p of EP)×(p of OL) Type of lens P of OL P of EP Total M OR Scanning 4x 10x 40x M= R of eye ÷ R of microscope Low power 10x 10x 100x Magnification Resolution Resolution of High power 40x 10x 400x of eye microscope Oil immersion 100x 10x 1000x Magnification power of objective lens ‫(الرقم اللي فوق‬ Numerical )‫على الشمال‬ aperture (NA) ‫(الرقم اللي فوق على‬ )‫اليمين‬ Distance between the objective and Thickness of ocular lenses coverslip ‫(الرقم اللي تحت على‬ ‫(الرقم اللي تحت على‬ )‫الشمال‬ )‫اليمين‬ Example 1: suppose that the maximum numerical aperture for oil immersion lens lenses is 1.5, calculate the maximum possible resolution of the light microscope? λ.0.6 0.527 × 0.6 R= = = 0.211 NA 1.5 The maximum possible resolution of the light microscope is 0.211 Example 2: if the resolving power of your eye (R) is 200 µm, what is the final magnification to see with the following objective lens? Numerical aperture from lens = 0.25 λ.0.6 0.527 × 0.6 Resolution of microscope = = = 1.265 NA 0.25 M = R of eye ÷ R of microscope = 200 ÷ 1.265 = 754.72 The final magnification of this objective lens is 754.72  Measurement of samples 1) Using of the micrometer slide to measure the size of specimen under the light microscope. 2) Using of scale bar to measure the size of photographed specimens (on Desktop by using different applications). Scale bar Micrometer slide Scale bar  BLOOD FILM (BLOOD SMEAR) What is blood smear ? It is a blood test to examine the abnormalities in blood cells. Red blood corpuscles or cells (RBCs) RBCs carry oxygen throughout human body THREE COMPONENTS White blood cells (WBCs) OF HUMAN WBCs help human body to fight infections BLOOD Platelets (PLTs) PLTs are very important for blood clotting  Steps of blood smear Preparation of Fixation of Staining of blood smear blood smear blood smear Preparation of blood OR smear Collection of Put in EDTA Finger stick blood peripheral blood (anticoagulant) directly onto slide Using within only one hour to avoid distortion of cells Equipment OR 1) Spreaders 2) Clean slides 3) Capillary tubes 3) Micropipette  1- place blood drop (2mm in diameter) approximately an inch from the frosted area of the slide 2- place the slide on a flat surface, hold the narrow side of the non frosted edge between your left thumb and forefinger 3- with your right hand, place the smooth clean edge of a second (spreader) slide on the specimen slide, just in front of the blood drop 4- allow the blood to spread almost to the edges of the second slide (or the spreader slide) 5- push the spread forward with one light, smooth and fluid motion. thin film of blood in the shape of bullet with a feathered edge will remain on the slide 6- label the frosted edge of the slide with name of patient, ID and the date of the blood test 7- allow the blood film to air-dry completely before staining (don’t blow to dry, the moisture from your breath will cause RBCs artifact)  To keep the morphology of cells, films must be fixed as soon as possible after they have dried Fixation of blood smear The best choice for fixation is methyl alcohol (methanol) as it is a good fixative To fix the films, put them in a covered staining jar or tray containing the alcohol for 2-3 minutes  Leishman stain Staining of JSB Giemsa blood smear stain stain stains Jenner Wright stain stain Field stain  Pour stain on slide drop by drop and wait 2 minutes Leishman Rinse the slide with water for 1-2 minutes stain Dry in air and examine under microscope with oil- immersion lens Methylene blue (basic ) stains acidic nucleus Leishman stain composed from Eosin (acidic) stains basic cytoplasm