Part II: Microscopy
quality control

Quality assurance with regard to tuberculosis microscopy is a system designed to continuously improve the reliability, efficiency and use of microscopy as diagnostic and monitoring option. The purpose of a quality assurance programme is to improve the efficiency and reliability of smear microscopy services.

The components of a quality assurance programme are

• quality control
• quality improvement
• proficiency testing

The following section will focus on aspects of quality control in the microscopy laboratory. For a discussion on quality improvement and proficiency testing please refer to the Management Series.

Quality control of microscopy is a process of effective and systematic internal monitoring of the performance of bench work in the microscopy laboratory. Quality control ensures that the information generated by the laboratory is accurate, reliable and reproducible. This is accomplished by assessing the quality of specimens, by monitoring the performance of microscopy procedures, reagents and equipment against established limits, by reviewing microscopy results and by documenting the validity of microscopy methods.

Quality control should be performed on a regular basis in the microscopy laboratory to ensure reliability and reproducibility of laboratory results. For a quality control programme to be of value, it must be practical and workable.

Quality control must be applied to:

• laboratory arrangement
• equipment
• collection and transport of specimens
• handling of specimens
• reagents and methods
• reporting of results

The keys to successful quality control are:

• adequately trained, interested and committed staff
• common-sense use of practical procedures
• a willingness to admit and rectify mistakes
• effective communication

Quality control measures which must be in place in all tuberculosis microscopy laboratories include:

Laboratory arrangement and administration

• Ensure that doors in the laboratory are always closed. Work areas, equipment and supplies should be arranged for logical and efficient work flow. Work areas should be kept free of dust. Benches should be swabbed at least once a day with an appropriate  disinfectant (eg. 5% phenol)
• Every procedure performed in the laboratory must be written out exactly as carried out and be kept in the laboratory for easy reference. Any changes must be dated and initialised by the laboratory supervisor
• All records should be retained for two years
• Laboratory procedures used routinely should be those that have been published in  reputable microbiological books, manuals or journals

Laboratory equipment

• Equipment should meet the manufacturers claims and specifications
• Written operating and cleaning instructions must be kept in a file for all equipment
• Dated service records must be kept for all equipment
• Equipment must be monitored regularly to ensure the constant accuracy and precision necessary. For the microscope this entails the following:

After daily use

• Wipe oil from objective, condenser and stage with lens paper soaked in xylene
• Turn off the microscope light source. Adjust the variable voltage regulator setting to minimum before turning off the lamp
• Replace microscope cover

Monthly

• Use an airbrush to blow away dust. (A simple airbrush can be made by attaching a Pasteur  pipette to a rubber bulb). Clean objectives, eye-pieces and condenser with lens paper soaked in xylene
• Remove slide holder from the stage and clean
• Wipe dust off the body of the microscope and the window of the illuminator in the base  using a water-moistened paper towel

Six-monthly

• Have the microscope  inspected, cleaned and lubricated by professional service personnel

Specimens and request forms

• Perform microscopy only upon written request of authorised persons and do not allow oral  requests without follow-up written instructions
• Insist on specimen request forms being kept separate from the specimens themselves.  Forms that have been contaminated by specimens should be sterilised by autoclaving or burnt
• Insist on adequately completed request forms and proper labelling of specimens to ensure positive identification of patients. Reject specimens that cannot be properly identified
• Evaluate the quality of sputum specimens and make a note if a specimen resembles saliva. The report should state ?specimen resembled saliva - treat a negative result with caution? (to facilitate reporting a rubber stamp with the comment can be made)
• Discard leaking and broken specimen containers by autoclaving and request a repeat specimen
• Document the arrival time of specimens in the laboratory and note any delays in delivery on the report form, particularly with negative results

Reagents and stains

• All containers of stains and reagents should show the date received and the date first  opened. Any material found to be unsatisfactory should be recorded as such and removed  from the laboratory immediately
• Stocks should be limited to six months? supply and regular stock rotation should take place to avoid unnecessary expiry

Staining and smear examination

• Stain slides in batches with a maximum of 12 slides per batch
• Include positive and negative controls with each day?s reading, especially if fewer than 10 slides are examined per day
• Read control slides before patient smears are read. Unacceptable control slides include the following:

Carbolfuchsin

• positive control is not stained red
• negative control remains red after decolourisation
• background is not properly decolourised

Fluorochrome

• negative control fluoresces
• positive control does not fluoresce or is dull
• background is not properly decolourised or fluoresces
• When the problem has been resolved, re-stain control slides as well as all patient  slides from the problem run
• Clean the slides with xylene and preserve them in separate slide boxes for external quality control. Pour 2-3ml of xylene onto the stained side of the slide and allow to dry.  Do not clean too vigorously or the stain may come off
• Retain all positive and negative slides, in the order in which the slides have been examined, for external quality control according to established procedures (see I.  Organisation and management).

Reporting and administration

• Send microscopy results out as soon as they become available and preferably within 24 hours of receipt of the sputum specimen
• Analyse microscopy results on a weekly and monthly basis for the percentage of positive results. Investigate any sharp differences from the norm. To identify excessive numbers of  positive results, compare the number of suspects examined to the number of smear-positives  identified: on average, for each smear-positive patient found there should be around 10 suspects examined. If there is a marked difference there may be a problem with the quality of microscopy or health workers may not be identifying suspects properly. The suspect to  case ratio of 10:1 may vary considerably from setting and it is recommended that the  average ratio be determined locally. Any deviation from the norm should then be  investigated
• Always check multiple positive smear results following on each other. This may suggest  transfer of organisms during smear preparation or during staining.