What happens if you leave a stain on a bacterial smear too long?

. What are some consequences of leaving a stain on a bacterial smear too long (over-staining)? Consequences of over-staining are that the cell wall may be broken up or completely destroyed which would result in a loss of morphological characteristics of the bacterial cell.

What happens if the stain is left on the bacterial smear too long?

Answer and Explanation: The consequences of over-staining the bacterial smear typically result in the loss of the distinct morphological features of the bacterial cells and...

What happens if you leave Gram stain on too long?

If the decolorizer is left on too long, even gram positive cells will lose the crystal violet and will stain red.

What happens if you leave a stain on a bacterial smear Not long enough?

If you leave the decolorizer on too long: gram positive bacteria will come out pink. If you don't leave it on long enough: gram negative bacteria will come out purple.

What is the consequence of having too much bacteria when performing a staining process?

If there are too many bacteria on the slide they will form a big glob and you will not be able to see the morphology of the individual cells. Large blobs of cells also do not stain properly and could yield erroneous results from the improper staining.

The Bacterial Smear

Why is time an important factor in simple staining?

Why is time an important factor in simple staining? Time is important because it creates a contrast between the bacteria and the stain. If you over or under stain you won't be able to see bacteria.

What happens when a bacterial smear is overheated?

If the smear is overheated during heat fixing, the cell walls will rupture. Concentration and freshness of reagents may affect the quality of the stain. Washing and drying of the smear between steps should be consistent. Excess water left on the slide will dilute reagents, particularly Gram's iodine.

What are the common errors in staining methods?

What may be the effects of the over decolorization on the results of the Gram staining method?

Over-decolorizing will lead to an erroneous result where gram-positive cells may stain pink to red indicating a gram-negative result, and under-decolorizing will lead to an erroneous result where gram-negative cells may appear blue to purple indicating a gram-positive result.

Why is it important to fix bacterial smears on a slide?

The purpose of making a smear is to fix the bacteria onto the slide. Fixing the bacteria will preserve the morphology of the cells long-term. Also, fixation assists the cells in adhering to the slide, so that the cells do not fall off the slide during the staining procedure.

What might cause a false gram-positive result?

Inappropriate specimen sampling, specimen processing, smear preparation, and prior antibiotic therapy are all factors that can have an adverse impact on Gram stain result. The inherent nature of some organisms may also produce misleading results; for example, Acinetobacter spp.

Why is the Gram stain not always successful?

Old cultures tend to lose the peptidoglycan cell walls, which predisposes gram-positive cells to be gram-negative or gram variable. Gram stain is not useful for organisms without a cell wall like Mycoplasma species, and for smaller bacteria like Chlamydia and Rickettsia species.

What would you observe if you Decolorized your slide too much?

What would you observe if you decolorized your slide too much? How would your cells appear? All of your cells would appear pink if you decolorized your cells too much with acetone-alcohol. Acetone-alcohol, if used too liberally, will wash the crystal violet out of the cells in your smear.

Why is an 18 24 hour culture necessary for a Gram stain?

Old cultures or smears could give atypical results. That is why cultures of 18-24 hours or recent smears are recommended. It is very important to control the heat-fixation (few seconds), any excess heating could produce erroneous results. Highly chlorinated tap water could weak the counter staining.

Why is it important to fix the smear quizlet?

The purpose of heat-fixing a smear is to make the cells stick to the slide. What is the purpose of heat-fixing a smear? Cell size, morphology (shape) and arrangement can be observed with simple stains.

What conditions can cause deceptive or incorrect Gram stain results?

Inappropriate specimen sampling, specimen processing, smear preparation, and prior antibiotic therapy are all factors that can have an adverse impact on Gram stain result. The inherent nature of some organisms may also produce misleading results; for example, Acinetobacter spp.

Which step is most likely to cause poor results in the Gram stain?

Excessive Decolorization

It is clear that the decolorization step is the one most likely to cause problems in the Gram stain.

Which factors can affect the results of a Gram stain and why?

The many variables that can affect this stain are age of the culture, amount of decolorizer used, the time of decolorization, the type of organism (acid-fast bacteria and spores do not stain well), thickness of the smear, and the general care of the stainer.

Why is it important not to overheat a bacterial smear?

Why should care be exercised not to overheat a bacterial smear when heat fixing to a slide? Overheating the slide can cause gas to expand in the slide and shatter it, presenting a physical danger. It will damage the cell wall and could lead to inconclusive results due to poor stain retention.

How would the pH of the stain affect the staining of bacteria?

How would the pH of the stain affect the staining of bacteria? Bacteria is slightly negative so it will attract the methylene blue as a basic stain and it will repel it as an acidic stain. Therefore, the basic stain would result in direct staining and the acidic stain would result in negative staining.

What is the most common source of errors during the Gram stain procedure?

In this study, we present a review of over 6,000 Gram-stains and establish an error rate of around 3%, with the most common reason for error being an over-decolourisation step resulting in organisms that should be Gram-positive appearing as Gram-negative.

What causes a stain to adhere to bacterial cells?

What causes a stain to adhere to bacterial cells? The positive charge of the edge will adhere to the negative charged cell surface structures.

Why should you be careful not to overheat the smear during the heat fixing step?

Why is it essential that smears be air-dried? Why can't they be gently heated over the flame to speed up the drying process? * Overheat will denature and rupture the cell wall. * Excess water on the slides will boil during fixation.

What would you expect from a Gram stain done on a slide that was heated too hot during the heat fixed smear?

The material must be fixed to the slide to prevent it from washing off during staining. This can be done by quickly passing the slide over a gentle flame (the slide should not become to hot to touch) or on a slide warmer. Overheating may alter cell morphology or cause organisms to decolorize more quickly.

What are the common mistakes in preparing bacterial smear?