Why are specimens to be stained suspended in water

It is easy to prepare distilled water at home. Simply fill a large pot with water and place a collection container inside the pot. As the water boils, the steam is collected inside the collection container.

This collected steam is distilled water and it should be stored in a sterile container. Just make sure it is a medical grade sterile water. Don't use boiled tap water. Ideally you should use propylene glycol PG as well. So if you wanted mL of solution, you would mix 1mL of trisodium citrate with 99mL of distilled sterile water.

Water in pristine form, as in the rain water or water obtained from snow is called pure water. When water is taken from other sources that is other than rain water it may contain salts and other dissolved solids and this is therefore impure water.

To remove these salts or other dissolved solid etc from the impure water the process of distillation is used. Distillation is a process in which water is boiled and heated till it evaporates and the evaporated vapour is cooled, thereby condenses to form water again.

This processed water is called distilled water. Uses of distilled water are for working, cleaning of sensitive chemical plants, machines etc, and in preparation of chemical solutions where pathogens, microorganisms are not of concern. It be noted that distilled water is pure water free from other chemicals etc but is not sterile. Sterile water is that which inhibits the growth of microorganisms. Thus the distilled water though pure, is not sterile water. So further processing of distilled water is done to make this water sterile.

Thus, the difference between distilled water and sterile water is that whereas distilled water is not free from microorganisms and may permit their growth, sterile water is totally free from microorganisms. No organic growth is possible in this. It is this sterile water which is used for injections and parentaral preparations in pharmacy.

The person's tongue is held down with a tongue depressor, as a healthcare worker moves a sterile, cotton-tipped swab across the back of the throat and tonsil region. If a sterile field becomes "contaminated" with a sterile solution, the field remains sterile.

Abscesses or fluids can be aspirated using a sterile syringe that is then tightly capped to prevent entry of air. The physician inserts a moistened, nonlubricated vaginal speculum.

After the cervix is exposed, the physician removes the cervical mucus. Next, he or she inserts a sterile cotton-tipped swab into the endocervical canal and rotates the swab. This definition is only one or the other. Either sterile or non-sterile. There is no such thing as partially sterile.

No it's not sterile. Log in. Study now. See Answer. Best Answer. Study guides. Biology 20 cards. There are no hard and fast rules for staining and rinsing times. The times listed are suggestions that usually work well. You will need to experiment with what works for the bacteria you have and the techniques you use. It is essential that you record exactly what you do and the results you observe in your lab book.

It would be useful for each lab bench member to pick a different stain so you can see what they all look like. Negative stains are even simpler than simple stains because you do not have to make a smear. A drop of cells is spread on a slide and viewed without fixation. The stain is a suspension of carbon, found in India ink or nigrosin. The carbon particles are negatively-charged, as is the cell membrane. The background looks black or sepia colored and the cells remain clear, since they repel the dye.

Some positively charged inclusion bodies, such as sulfur, may stain. This stain gives accurate information on cell morphology and capsule presence because the cells are not fixed. Cell size appears slightly larger because any extracellular coatings or secretions on the outside of the cell membrane also do not stain.

Negative stains are useful for rapid determination of the presence of Cryptococcus neformans , the causative agent of cryptococcisis, in cerebral spinal fluid. This technique is also used when you stain for endospores and capsules. Just as in preparing a smear, you only need a small amount of organism. It is also important not use too much nigrosin. If it is too thick, the background will have a cracked appearance similar to mud puddles drying in the sun.

You want to get a light film. Your instructor will demonstrate this technique for you. Nigrosin comes off the slide and onto your oil immersion lens very easily.

Be sure to thoroughly clean your oil lens when you are finished. Then clean it again. Once it dries on the lens it is very difficult to remove and will impair your ability and the other micro students using that scope to see clearly out of the lens. The Gram stain is the most common differential stain used in microbiology.

Differential stains use more than one dye. The unique cellular components of the bacteria will determine how they will react to the different dyes. The Gram stain procedure has been basically unchanged since it was first developed in Almost all bacteria can be divided into two groups, Gram negative or Gram positive. A few bacteria are gram variable. Trichomonas , Strongyloides , some fungi, and some protozoa cysts also have a Gram reaction.

Very small bacteria or bacteria without a cell wall, such as Treponema , Mycoplasma , Chlamydia , or Rickettsia do not have a gram reaction. The characterization of any new bacteria must include their gram reaction. Typically a differential stain has four components; the primary stain, a mordant that sets the stain, a decolorizing agent to remove the primary stain, and a counter stain.

In the Gram stain, the primary stain is crystal violet. This gives the cell an intense purple color. The mordant, iodine, forms a complex with the crystal violet inside the cell wall.

Gram positive cells will retain the dye complex and remain purple. The dye rinses out in gram negative cells. The large iodine-crystal violet complex is retained within the cell walls of gram positive cells because of the molecular structure of the many layers of peptidoglycan in the cell wall. There are lots of cross-linked teichoic acids and the iodine-dye complex cannot physically get out. There is also less lipid in the membrane and the decolorizing agent cannot get to it as well.

Gram negative cells have an outer membrane and only one layer of peptidoglycan, with more lipid. The crystal violet dye is easily washed out. The accuracy of the Gram stain is dependent on the integrity of the bacterial cell wall.

There are a variety of things that can influence the cell wall integrity; old cells i. Under these conditions, gram positive cells will come out as gram-negative.

If you de-colorize too long, Gram-positive cells will look like Gram-negative cells. Conversely, if you do not decolorize enough, Gram-negatives will look like Gram-positives.

The only way you can trust your results it to always run a known Gram-positive and a known Gram-negative on the same slide. If they stain as predicted you can be pretty sure the result of your unknown sample is reliable.

The Gram staining takes practice to get right. Do not expect to get a good Gram stain on your first try. Endospore-forming bacilli note endospores as small, round, hollow, unstained areas, within or at one end of bacillary bodies ; 4.

A bacillus showing pleomorphism note varying widths and lengths. Spirilla short curved or spiraled forms with rigid bodies ; 2. Spirochetes long tightly or loosely coiled forms with sinuous flexible bodies.

Some may show great variation in their size and length pleomorphism. Spiraled bacteria occur singly and usually do not form group patterns. Examine color-plates 1—8 to see representative examples of bacterial morphology. Slides for microscopic smears must always be sparkling clean. They may be stored or dipped in alcohol and polished clean free of grease with a tissue or soft cloth. At one end of the slide write the initials of one of the three assigned organisms your three slides should read Se, Bs, and Ec, respectively.

Turn the slides over so that the unmarked side is up. When slides are to be stained, pen or pencil markings should always be placed on the underside so that the mark will not smear, wash off, or run into the smear itself. With your inoculating loop, place a loopful of water in the ringed area of the slide. Using proper aseptic transfer techniques, mix a small amount of bacteria in the water and spread it out.

Repeat this step until smears of all three organisms have been made. Allow the smears to air dry. You should be able to see a thin white film on each slide. If not, add another loopful of water and more bacteria as in step 4.

Heat-fix the smears by passing the slides rapidly through the Bunsen flame three times so that the smears will not wash off.

If a Bunsen burner is not available, fix the smears by placing the slides on a staining rack and flooding them with absolute methanol. Allow the slides to sit for one minute, then drain off the alcohol and air dry them completely. Place the slides on a staining rack and flood them with methylene blue. Leave the stain on for three minutes. Wash each slide gently with distilled water, drain off excess water, blot do not rub with bibulous paper, and let the slides dry completely in air.

While the slides are drying, take two more clean slides and draw a circle on the bottom with your wax pencil or marking pen. With the flat end of a toothpick, scrape some material from the surface of your teeth and around the gums. Emulsify the material in the drop of water on one slide. Repeat this procedure on the other slide.

Allow both slides to dry in air; then fix them with heat or methanol. Stain one slide with methylene blue for three minutes and the other with safranin for three minutes.