Laboratory Medicine and Diagnostic Pathology

Laboratory Medicine and Diagnostic Pathology

Brian C. Muzyka, DMD, MS, MBA

John Christie, MD, PhD

Bobby Collins, DDS, MS

Pathology has traditionally been divided into laboratory medicine and anatomic pathology. Laboratory medicine is the division of medical science dealing with the qualitative and quantitative assessment or analysis of blood (formed elements and fluid components) and other cells and body fluids, while anatomic pathology involves the determination of disease through gross and microscopic examination of tissue. Clinical chemistry, microbiology, hematology, immunology, molecular diagnostics, cytology, toxicology, therapeutic drug monitoring, genetics, and histology are various components of laboratory medicine.

Diagnostic testing is performed on samples of blood, tissue, and other cells and bodily fluids to provide the clinician with information about a person’s health, such as establishing a diagnosis of a condition or disease; screening for disease in asymptomatic patients; and excluding the diagnosis of a disease or condition or monitoring therapy.

The need for this chapter on laboratory medicine in a textbook of oral medicine stems from the National Academy of Medicine report in 2015, which indicated that the medical history of almost every adult in the United States contains a minimum of one diagnostic error.1 It is difficult to determine the impact of laboratory testing on medical decisions. Historically it has been accepted that 70% of medical decisions are a direct result of laboratory testing.1 More recently, reports of the use of laboratory tests in patient care medical decisions ranged from 38% for general internists and 29% for family physicians.2 The percentage of medical decisions made as a direct result of laboratory results is apparently less than the historically estimated 70%, but clearly laboratory testing plays a major role in the provision of healthcare. The Centers for Medicare and Medicaid Services (CMS) estimate that the overall spending on healthcare in the United States in 2017 was $3.5 trillion. National healthcare spending in the United States is projected to grow at an average rate of 5.5% per year for 2018–2027 and to reach nearly $6.0 trillion by 2027.3 Laboratory testing accounts for approximately $97 billion, 28% of overall US healthcare spending per year. In 2027, the estimated economic impact of laboratory testing in the United States will approach $1.68 trillion.

A significant proportion of diagnostic errors in medical histories are due to problems with ordering, performing, reporting, and interpreting laboratory tests. Unfortunately, schools of medicine, dentistry, and allied health do not routinely provide formal courses in laboratory medicine. With the increasing complexity of laboratory testing and little clinically oriented support from practitioners of laboratory medicine, medical, dental, pharmacy, and nursing professionals are confused and uncertain about the proper use of laboratory results.

The aims of this chapter are to provide an introduction to the general principles of laboratory medicine, which should be a part of every oral healthcare professional’s knowledge base, as well as help fulfill two of the goals of the Institute of Medicine report: to employ more effective teamwork in the diagnostic process and to enhance healthcare professionals’ education and training in the diagnostic process (Table 28‐1).

Laboratory testing includes three steps: preanalytic—activities such as test ordering and sample collection that take place before actual testing; analytic—laboratory performance of the test; and postanalytic—the process of reporting and interpreting the result. As the analytic phase of laboratory testing is highly regulated and under tight quality control, most laboratory‐associated errors occur in the preanalytic and postanalytic stages. For this reason and also for the reason that those two areas are most affected by the decisions of medical/dental professionals, this chapter will focus on the preanalytic and postanalytic stages of laboratory testing.


Appropriate laboratory testing is defined as “ordering of the right test, using the right method, at the right time, to the right patient, with the right costs and producing the right outcome.”4 The increasing emphasis on the appropriateness of laboratory testing is shown by the development of initiatives such as the “Choosing Wisely” campaign (, developed by the American Board of Medicine, and laboratory stewardship programs such as PLUGS (Patient‐centered Laboratory Utilization Guidance Service), started by Seattle Children’s Hospital and found in many hospitals (,6

Table 28‐1 Institute of medicine september 2015 diagnosis recommendations.

Source: Institute of Medicine (2015) Recommendations: improving diagnosis in health care. Washington, DC: National Academies of Sciences, Engineering, Medicine.

Goal 1: Facilitate more effective teamwork in the diagnostic process among health care professionals, patients, and their families
Recommendation 1A
In recognition that the diagnostic process is a dynamic team‐based activity, health care organizations should ensure that health care professionals have the appropriate knowledge, skills, resources, and support to engage in teamwork in the diagnostic process. To accomplish this, they should facilitate and support:

  • Interprofessional and intraprofessional teamwork in the diagnostic process.
  • Collaboration among pathologists, radiologists, other diagnosticians, and treating health care professionals to improve diagnostic testing processes.
Recommendation 1B
Health care professionals and organizations should partner with patients and their families as diagnostic team members and facilitate patient and family engagement in the diagnostic process, aligned with their needs, values, and preferences. To accomplish this, they should:

  • Provide patients with opportunities to learn about the diagnostic process.
  • Create environments in which patients and their families are comfortable engaging in the diagnostic process and sharing feedback and concerns about diagnostic errors and near misses.
  • Ensure patient access to electronic health records (EHRs), including clinical notes and diagnostic testing results, to facilitate patient engagement in the diagnostic process and patient review of health records for accuracy.
  • Identify opportunities to include patients and their families in efforts to improve the diagnostic process by learning from diagnostic errors and near misses.
Goal 2: Enhance health care professional education and training in the diagnostic process
Recommendation 2A
Educators should ensure that curricula and training programs across the career trajectory:

  • Address performance in the diagnostic process, including areas such as clinical reasoning; teamwork; communication with patients, their families, and other health care professionals; appropriate use of diagnostic tests and the application of these results on subsequent decision making; and use of health information technology (IT).
  • Employ educational approaches that are aligned with evidence from the learning sciences.
Recommendation 2B
Health care professional certification and accreditation organizations should ensure that health care professionals have and maintain the competencies needed for effective performance in the diagnostic process, including the areas listed in Recommendation 2A.
Goal 3: Ensure that health information technologies support patients and health care professionals in the diagnostic process
Recommendation 3A
Health IT vendors and the Office of the National Coordinator for Health Information Technology (ONC) should work together with users to ensure that health IT used in the diagnostic process demonstrates usability, incorporates human factors knowledge, integrates measurement capability, fits well within clinical workflow, provides clinical decision support, and facilitates the timely flow of information among patients and health care professionals involved in the diagnostic process.
Recommendation 3B
ONC should require health IT vendors to meet standards for interoperability among different health IT systems to support effective, efficient, and structured flow of patient information across care settings to facilitate the diagnostic process by 2018.
Recommendation 3C
The Secretary of the U.S. Department of Health and Human Services (HHS) should require health IT vendors to:

  • Routinely submit their products for independent evaluation and notify users about potential adverse effects on the diagnostic process related to the use of their products.
  • Permit and support the free exchange of information about real‐time user experiences with health IT design and implementation that adversely affect the diagnostic process.
Goal 4: Develop and deploy approaches to identify, learn from, and reduce diagnostic errors and near misses in clinical practice
Recommendation 4A

  • Accreditation organizations and the Medicare conditions of participation should require that health care organizations have programs in place to monitor the diagnostic process and identify, learn from, and reduce diagnostic errors and near misses in a timely fashion. Proven approaches should be incorporated into updates of these requirements.
Recommendation 4B
Health care organizations should:

  • Monitor the diagnostic process and identify, learn from, and reduce diagnostic errors and near misses as a component of their research, quality improvement, and patient safety programs.
  • Implement procedures and practices to provide systematic feedback on diagnostic performance to individual health care professionals, care teams, and clinical and organizational leaders.
Recommendation 4C

  • HHS should provide funding for a designated subset of health care systems to conduct routine postmortem examinations on a representative sample of patient deaths.
Recommendation 4D

  • Health care professional societies should identify opportunities to improve accurate and timely diagnoses and reduce diagnostic errors in their specialties.
Goal 5: Establish a work system and culture that supports the diagnostic process and improvements in diagnostic performance
Recommendation 5
Health care organizations should:

  • Adopt policies and practices that promote a non‐punitive culture that values open discussion and feedback on diagnostic performance.
  • Design the work system in which the diagnostic process occurs to support the work and activities of patients, their families, and health care professionals and to facilitate accurate and timely diagnoses.
  • Develop and implement processes to ensure effective and timely communication between diagnostic testing health care professionals and treating health care professionals across all health care delivery settings.
Goal 6: Develop a reporting environment and medical liability system that facilitates improved diagnosis by learning from diagnostic errors and near misses
Recommendation 6A
The Agency for Healthcare Research and Quality (AHRQ) or other appropriate agencies or independent entities should encourage and facilitate the voluntary reporting of diagnostic errors and near misses.
Recommendation 6B
AHRQ should evaluate the effectiveness of patient safety organizations (PSOs) as a major mechanism for voluntary reporting and learning from these events and modify the PSO common formats for reporting of patient safety events to include diagnostic errors and near misses.
Recommendation 6C
States, in collaboration with other stakeholders (health care organizations, professional liability insurance carriers, state and federal policy makers, patient advocacy groups, and medical malpractice plaintiff and defense attorneys), should promote a legal environment that facilitates the timely identification, disclosure, and learning from diagnostic errors. Specifically, they should:

  • Encourage the adoption of communication and resolution programs (CRPs) with legal protections for disclosures and apologies under state laws.
  • Conduct demonstration projects of alternative approaches to the resolution of medical injuries, including administrative health courts and safe harbors for adherence to evidenced‐based clinical practice guidelines.
Recommendation 6D
Professional liability insurance carriers and captive insurers should collaborate with health care professionals on opportunities to improve diagnostic performance through education, training, and practice improvement approaches and increase participation in such programs.
Goal 7: Design a payment and care delivery environment that supports the diagnostic process
Recommendation 7A
As long as fee schedules remain a predominant mechanism for determining clinician payment, the Centers for Medicare & Medicaid Services (CMS) and other payers should:

  • Create current procedural terminology (CPT) codes and provide coverage for additional evaluation and management activities not currently coded or covered, including time spent by pathologists, radiologists, and other clinicians in advising ordering clinicians on the selection, use, and interpretation of diagnostic testing for specific patients.
  • Reorient relative value fees to more appropriately value the time spent with patients in evaluation and management activities.
  • Modify documentation guidelines for evaluation and management services to improve the accuracy of information in the EHR and to support decision making in the diagnostic process.
Recommendation 7B
CMS and other payers should assess the impact of payment and care delivery models on the diagnostic process, the occurrence of diagnostic errors, and learning from these errors.
Goal 8: Provide dedicated funding for research on the diagnostic process and diagnostic errors
Recommendation 8A
Federal agencies, including HHS, the U.S. Department of Veterans Affairs, and the United States Department of Defense, should:

  • Develop a coordinated research agenda on the diagnostic process and diagnostic errors by the end of 2016.
  • Commit dedicated funding to implementing this research agenda.
Recommendation 8B
The federal government should pursue and encourage opportunities for public–private partnerships among a broad range of stakeholders, such as the Patient‐Centered Outcomes Research Institute, foundations, the diagnostic testing and health IT industries, health care organizations, and professional liability insurers to support research on the diagnostic process and diagnostic errors.

In order to reduce or eliminate activities that have little benefit or can cause harm, Choosing Wisely focuses on what not to do rather than on positive advice about what to do. The recommendations from Choosing Wisely are helpful and, as a result, many medical specialty societies have adapted Choosing Wisely and include it as part of their medical specialty website.

In patient care, the first step in ordering the appropriate laboratory test is to determine whether a test should be ordered. In 1994, Dr. Catherine DeAngelis, the first woman editor of the Journal of the American Medical Association, made a pithy comment that summarizes the whole issue very succinctly: “Remember ordering a laboratory test is a bit like picking your nose in public: you must consider what you will do if you find something.”7 Hooper and his colleagues put the same sentiment in slightly different language: “Before ordering a test, decide what you will do if it is either positive or negative. If both answers are the same, then don’t do the test.“8

Lee in the 25th edition of Goodman‐Cecil Medicine had the following comments about laboratory testing that provide more detailed advice about ordering laboratory tests:

The interpretation of test results depends on what is already known about the patient.

No test is perfect. Clinicians should be familiar with their diagnostic performance and never believe that a test “forces” them to pursue a specific management strategy.

Tests should be ordered if they may provide additional information beyond that already available.

Tests should be ordered if there is a reasonable chance that the data will influence patient care.

Two tests that provide similar information should not be used.

In choosing between two tests that provide similar data, use the one that has lower cost and/or causes less discomfort and inconvenience to the patient.

Clinicians should seek all the information provided by the test, not just an abnormal or normal result.

The cost‐effectiveness of strategies using noninvasive tests should be considered in a manner similar to that of therapeutic strategies.9

Consideration of which laboratory test to order involves examination of parameters of the specific assay that determine the diagnostic usefulness of the test. In determining the usefulness of a test, the concepts of sensitivity and specificity come into play. In a population of patients in which the correct disease status is already known, sensitivity is the percentage testing positive in a population with disease (positive in disease), while specificity is the percentage testing negative in a population without disease (negative in health). For most commercially available assays, the sensitivity and specificity of a particular test are listed in the literature provided by the manufacturer. However, the sensitivity and specificity measures provided by the manufacturer are obtained under ideal conditions that are generally not found in clinical use. Generally, in “real‐life” situations, the actual sensitivity and specificity of a particular assay are less than those indicated in the manufacturer’s instructions for the test. As a general rule, a diagnostic assay should have performance characteristics that minimize false positives and false negatives. However, the clinical implications of a test result may determine which test is most useful in a specific situation. For patients who potentially have a serious, possibly incurable disease, the diagnostic test should have high specificity so as to minimize the number of false positive results. On the other hand, an assay that maximizes sensitivity should be used to diagnose serious but curable diseases. A second confirmatory assay may then be used.

When choosing between the sensitivity and specificity of a laboratory test for a condition, clinicians may find the mnemonic “SpIn and SnOut” helpful. Specificity, abbreviated as Sp, is used to rule in the condition (SpIn). Sensitivity, abbreviated as Sn, is used to rule out the condition (SnOut) (see Figure 28‐1).

The actual usefulness of a laboratory test is better predicted by the positive and negative predictive values. The positive predictive value (PPV) of a test is the percentage of all positive results that are true positives and is calculated using the formula:


The negative predictive value (NPV) is the percentage of all negative results that are true negatives and is calculated using the formula:


A good test should have both high positive and negative predictive values. The predictive values for any test depend not only on the sensitivity and specificity of the test, but also on the prevalence of the disease for the diagnostic laboratory assay.

Schematic illustration of mnemonic for Differentiating between Sensitivity and Specificity.

Figure 28‐1 Mnemonic for Differentiating between Sensitivity and Specificity.

Using systemic lupus erythematosus (SLE) and the antinuclear antibody (ANA) test as an example:

The sensitivity and specificity for ANA in the diagnosis of SLE are 95% and 90%, respectively; testing for SLE with ANA in all patients presenting to a primary care practice (assume prevalence of < ?span Start cssStyle="text-decoration:underline"?><1%), the PPV and NPV are 9% and >99%, respectively. In a rheumatology clinic in which only patients with clinical features of SLE (assume prevalence of 30%) are tested with ANA, the PPV and NPV of the test are 80% and 96%, respectively.

Understanding PPV and NPV is important for the dental healthcare team. Dental advertising and dental salespersons may try to sell tests for diagnosis of caries, periodontal disease, oral cancer, or other diseases and conditions to general dental practices. Moreover, these tests may be advertised for direct sale to the public. Whether the test is used for screening or diagnosis depends on the characteristics of the assay: high sensitivity for screening tests and high specificity for exclusion of a diagnosis. Preferably those characteristics should be obtained from published literature and not from the detail sheet accompanying the materials for performance of the assay. Dental practitioners using these tests need to be aware of the predictive value of these tests in their own patient populations and determine whether or not these tests are useful, and whether these assays are more appropriate for screening than for diagnostic purposes.

The diagnostic accuracy of a particular test is provided by knowledge of the sensitivity and specificity for that assay. However, these values do not give any indication about the effect the result has on the likelihood that the patient has the disease for which the test is performed. The relationship between the clinical suspicion—that is, the pre‐test probability—that a patient has a disease compared to the chance that the patient has the disease after a particular test is performed—that is, the post‐test probability—is given by the formula:


where LR is the likelihood ratio.

The pre‐test probability is the clinician’s estimate that the patient has the disease. This probability is estimated from the prevalence of the disease in the population from which the patient comes and the history and physical examination.

The LR of a positive result is calculated by the following equation:


The LR of a negative result is calculated by the following equation:


The diagnostic or testing threshold of a test is the point at which clinical suspicion for disease is so low that the results of the laboratory test will not alter that low probability. If the test has a low diagnostic threshold even if the test is positive, it is most likely a false positive result and the patient should not be treated for that disease. The therapeutic or treatment threshold is the point at which clinical suspicion is high enough that the results of the assay will not significantly increase the likelihood that the patient has the disease. Laboratory tests should be ordered when the pre‐test probability of the patient having the disease lies between the diagnostic and the therapeutic threshold (see Figure 28‐2). These thresholds are generally determined by published analyses. An example of determination of these break‐points is seen in two papers dealing with Lyme disease testing.10,11 In these papers, the diagnostic or testing threshold was 0.20 (a patient with nonspecific arthritis) and the therapeutic threshold was 0.8 (a patient from a geographic area endemic for Lyme disease with either erythema migrans or a combination of arthritis, history of a rash resembling erythema migrans, and a previous tick bite).

Diagnostic testing can be performed in a multitude of locations. Laboratories with a large menu of diagnostic tests generally include those laboratories found in larger hospitals, hospital systems, and commercial laboratories, such a Quest Diagnostics or LabCorp. Most hospitals and large healthcare centers have in‐house laboratories. These in‐house laboratories may offer a full spectrum of laboratory testing or may be limited to more routinely used assays, and the in‐house laboratory may send out less commonly used assays to a commercial laboratory. Diagnostic testing also occurs in smaller labs found in providers’ offices, healthcare centers, urgent care centers, or other locations such as pharmacies. In the outpatient nonhospital setting, specimens are generally obtained from patients and sent to a centralized regional lab for analysis.

Schematic illustration of diagnostic tree for ordering a test.

Figure 28‐2 Diagnostic tree for ordering a test.

In the United States, diagnostic laboratories are required to be licensed and certified. On October 31, 1988, the US Congress enacted the Clinical Laboratory Improvement Amendments of 1988 (Pub. L. 100‐578) (CLIA’88), codified at 42 U.S.C. 263.12 This act, more generally known as CLIA, established quality standards for laboratory testing to ensure the accuracy, reliability, and timeliness of patient test results regardless of the location of the testing site, and requires laboratories to be CLIA certified. CLIA certification is important, as a clinical laboratory must be CLIA certified to receive reimbursement from Medicare or Medicaid. In the United States, Medicare payments for lab tests totaled $6.8 billion in 2016 and reimbursement for services provided is certainly an incentive for laboratories to obtain and maintain their CLIA certification.

Point‐of‐Care Testing

Office laboratories often use point‐of‐care (POC) assays as the basis for much of their testing. Point‐of‐care testing (POCT), defined as any testing that occurs outside of a central laboratory, is becoming a more common and accepted practice in the healthcare industry. POCT offers several advantages over traditional, more regulated laboratory testing, including increased accessibility to diagnostic testing for many populations, especially in rural and remote settings; minimal sample volumes required to perform the assay; real‐time test results; and, most importantly, the ability of nonlaboratory personnel to perform these assays. However, the instructions for these POC tests must be followed precisely; and, as in more traditional laboratories, there must be documentation of training, performance of controls and instrument checks, and correct performance of procedures.

As useful as POC assays are, they may be inappropriate in certain clinical situations. An example is the use of a POC assay to determine glycosylated hemoglobin (HbA1c) for the diagnosis of diabetes mellitus. HbA1c is a measure of blood glucose over the life span of the red blood cell, which is approximately 120 days. While it may be appropriate to use POC HbA1c assays to guide outpatient treatment of patients with diabetes, the analytic performance of these tests—that is, their accuracy relative to a reference laboratory’s standard and precision—is such that patients who have true HbA1c values close to the diagnostic cut‐point may be misdiagnosed. A cut‐point is a value at which subjects are classified as having or not having a condition. The World Health Organization (WHO) recommends an HbA1c cut‐point of 6.5% for the diagnosis of diabetes. A value of less than 6.5% does not exclude a diabetes diagnosis and further clinical investigation may be required.13 Even in the same institution, there is often a difference between the results of an assay performed in an outpatient/clinic setting by one methodology and an assay for the same analyte performed by the reference method in the main hospital laboratory. When such differences are clinically important, such as in diagnosis of a disease, the main laboratory should append a comment to the results to indicate that such discrepancies exist and that correction factors have been applied to make the results compatible.

The American Dental Association (ADA) advocates for POCT of patient blood glucose levels on an ongoing basis or immediately prior to dental treatment as an appropriate activity for persons at risk for diabetes. In support of this recommendation, the ADA has established a billing code (D0142) for this test (Box 28.1).

Additionally, the ADA has established a billing code for testing for HbA1c (Box 28.2).

Further information on ADA recommendations for POC testing can be found on the ADA website.14

Laboratory Analysis

Laboratory analysis is based upon the idea that the material sampled will be collected in a standardized fashion to reduce the likelihood of undue variability of results. Since patients are not standardized, their physical status can significantly affect their hematologic profile and laboratory results. The patient’s degree of hydration, level of anxiety, activity, medications, history of tobacco use, sex, and race may impact upon hematologic profile results. Additionally, the presence of certain medical conditions may impact laboratory test results. For example, HbA1c values are elevated in iron‐deficiency anemia patients and decrease after the patients are treated with iron.15

Descriptive information regarding the patient is required when submitting a sample for laboratory analysis. This descriptive information may include pertinent medical history, sex, age, race, fasting status, and time of collection. Correctly labeling specimens that have been obtained is of the utmost importance. Samples must be labeled with appropriate identification to ensure the test results will be related back to the patient. The use of < ?span Start cssStyle="text-decoration:underline"?>>1 unique patient identifiers such as medical record numbers or birthdate along with bar codes generated to identify that a sample was obtained from a specific known patient has become standard practice in clinics and hospitals. The barcode and unique identifiers are placed on the specimen container and attached to the laboratory requisition form.

In collecting blood for determination of specific analytes, there are different types of collection tubes whose purpose is generally indicated by the color of the top of the tube. Table 28‐2 gives a representative sample of the common blood collection tubes and their uses.16,17 However, the laboratory to which you send samples should provide a list of the blood collection tubes, specimen requirements, and any directions for use. If you are unsure about any of this information, you should contact the laboratory directly.

In order to aid in the draw of blood into the container, most of these tubes contain a vacuum. The tubes should be filled with a certain amount of blood with at least the minimum specimen volume designated by the laboratory. The tops of the tubes should not be removed to completely fill the container. In tubes that contain additives, under‐ or overfilling can yield in false results. Many of the collection tubes with additives should be inverted several times to allow complete mixing of the blood and/or any additive. Order of draw, when obtaining multiple samples for testing, is important, with the following sequence being recommended: sterile tubes for blood culture; tube for coagulation testing (light blue); gel separator tube (red); tube with heparin additive (green tube); tube with EDTA additive (purple/lavender tube); tube with oxalate/fluoride additive (gray tube).

Falsely high or low results for assays can occur due to materials that interfere with the accurate and precise measurement of the analyte in question. As many laboratory results are determined by spectrophotometric methods, any substance that colors the plasma and serum can interfere with the results. The three major interferences due to abnormal colors in a liquid are lipemia, causing a milky‐white color; hemolysis, resulting in a red color; and increased bilirubin, turning plasma/serum orange, green, or brown. Besides altering the color of serum/plasma, many drugs produce significant interference by other means. Therefore, it is important to include medication information on a laboratory requisition slip when requested.

For most tests, blood can be drawn at any time during the day. However, there are exceptions to this statement. For therapeutic monitoring of drugs such as antibiotics, the medical professional needs to know if the drug concentration is within a certain therapeutic range. It is easier to draw a trough level, the lowest level of a drug before administration of the next dose, than a peak level for a drug. A peak level is the highest level of drug in the serum and is usually obtained 1 to several hours after the drug is administered. For an intravenously administered drug, blood is drawn for trough levels just prior (< ?span Start cssStyle="text-decoration:underline"?><30 minutes ideally) to administration of the next dose. Some biochemical assay results are biased by diurnal variations. Time of collection must be considered when interpreting these values in the clinical setting. For example, sampling of blood for distinction of pseudo‐Cushing states from Cushing syndrome gives 96% accuracy. In Cushing’s syndrome the loss of normal cortisol circadian rhythm and absence of a late‐night cortisol nadir is typical. Midnight serum cortisol levels have been used to distinguish patients with Cushing’s syndrome from those with pseudo‐Cushing’s syndrome. However, drawing blood at midnight is inconvenient for an ambulatory patient, so obtaining saliva for cortisol determination at bedtime or midnight is more convenient and just as accurate. For those readers who wish more information about laboratory medicine, including laboratory reference ranges, we recommend the third edition of Laposata’s Laboratory Medicine Diagnosis of Disease in the Clinical Laboratory.16

Table 28‐2 Selected examples of blood collection tubes.

Sources: Modified from Laposata M. Laposata’s Laboratory Medicine: Diagnosis of Disease in the Clinical Laboratory, 3rd edn. New York: McGraw‐Hill, 2019; Bakerman S, Bakerman P, Strausbauch P. Bakerman’s ABC’s of Interpretive Laboratory Data, 5th edn. Scottsdale, AZ: Interpretive Laboratory Data, 2014.

Cap Color Content(s) Comments and Uses
Red None Clotted blood or serum
Most routine chemistries,
blood bank, serology
Light blue Sodium citrate Acts as anticoagulant, plasma
coagulation testing
Purple (lavender) EDTA Acts as anticoagulant
Unclotted blood for
hematology, genetic testing,
immunosuppressants, red blood cell folate, HbA1c
Green Sodium/lithium Acts as anticoagulant
heparin heparinized plasma, whole
blood, and bone marrow
Gold Clot activator Gel for serum separation after
with gel centrifugation; shorter clot
Most routine chemistries
Do not use for toxicology or
drug testing
Gray Sodium fluoride Fluoride inhibits glycolysis,
+ EDTA or allowing optimum glucose
potassium oxalate measurements
EDTA and oxalate are
Yellow Acid‐citrate dextrose Anticoagulates blood and
solution preserves cells during
Blood bank studies, human leukocyte antigen
Dark blue EDTA or none Trace element determination

Although oral infections are common, bacterial cultures are of limited value in dentistry. Often, all that is needed is a direct smear of the infected tissue of interest, such as fluid from a vesicle, abscess aspirate, or scraping from tongue or mucosal sites. Microbiologic cultures of soft tissue infections of the head and neck are frequently not performed because there is a mixed aerobic/anaerobic flora that is difficult to sample and to culture. For these infections in which a culture has been submitted, most laboratories will either give a diagnosis of mixed aerobic/anaerobic flora and/or list the three most predominant organisms. However, for severe infections such as osteomyelitis, acute parotitis, and cellulitis, culture and sensitivity should be performed. If cultures are to be submitted to a diagnostic laboratory, pus or tissue is the best choice of sample materials. Microbiologic swabs are convenient, but inferior to tissue and fluid. Tissue and fluid are essential for fungal and mycobacterial cultures. Moreover, if there is a possibility that an anaerobic organism is the causative organism, materials for culture should be submitted in anaerobic transport containers. Unless the antibiotic susceptibility of a bacterium can be predicted from its identity, a standardized battery of antibiotic susceptibilities is performed as part of the culture. However, unless fungal and mycobacterial cultures are sent to a large reference laboratory, often susceptibilities will only be performed with specific antifungal or antimycobacterial drugs upon specific request.

In the clinical pathology area, molecular methods have probably made their greatest inroads in the microbiology laboratory. In many virology reference laboratories, molecular methods with their greater sensitivity have replaced viral cultures. Many companies are marketing polymerase chain reaction (PCR) tests that can be performed in small laboratories. Many of these kits can detect multiple organisms (multiplex) from one sample. However, there is a growing awareness that detecting several organisms in one sample means that the clinical significance of these results is often hard to determine. In quantitative PCR assays, used most frequently for determination of viral loads in blood or tissue, the variability is such that a log10 difference between two different samples is necessary before it can be assumed that there is a true difference between results. In addition, the results of two different quantitative PCR tests performed in different laboratories cannot be compared.

Infectious diseases are one area in which correlation of laboratory results—that is, those from the microbiology laboratory with those from tissue biopsies of the same site—is absolutely essential. With increasing degrees of immunosuppression and more powerful drugs, it is now recognized that organisms that were formerly considered colonizers or of low virulence can invade tissues and be pathogenic. This is especially true of the majority of fungi, which are considered opportunistic pathogens. For example, sinus material from patients with chronic sinusitis often grows Aspergillus spp. In immunosuppressed patients the finding of Aspergillus spp. does not help the clinician determine if the diagnosis is chronic fungal sinusitis or invasive fungal sinusitis with the accompanying possibility of dissemination to other sites. Only a tissue biopsy will provide definitive evidence of sinus fungal invasion. There are other situations in which culture will provide a definitive identification of an organism, but tissue is necessary for determination of the clinical significance of the organism. Examples include active Candida spp. infection from a smear of involved epithelium demonstrating the presence of hyphae; the presence of characteristic viral inclusions in tissues for diagnosis of viral invasion; and the presence of the agents of endemic mycoses in histologic/cytologic material, as cultures may take weeks to grow or fail to grow.

For those organisms that cannot be detected by the methods we have previously discussed, serologic methods are often valuable. These methods detect the presence of a specific antigen in a body fluid or the presence of specific antibodies directed against the organism. Unfortunately, for many exotic pathogens, serologic diagnosis depends on assays available only in large, reference laboratories or at facilities such as the Centers for Disease Control and Prevention (CDC).

Theoretically, the most specific methods for measuring antibody production detected against a particular pathogen depend on the detection of immunoglobulin (Ig) M for documentation of an acute infection or measurement of an antibody, usually IgG, at two different time intervals, weeks apart. In many cases, these measures are not practical: either the peak of IgM has passed, or it is unlikely that the patient will return for a second visit to the laboratory for a blood draw weeks distant for the initial clinic visit. In such cases, the clinician must depend on an assay that detects the absolute magnitude of the antibody response at the time of the clinic visit.

Healthcare providers may order laboratory tests by completing an appropriate medical order for the test. Many of the major laboratories have online systems to request testing and to view the results. In order to use these online systems, the clinician must register with these laboratories. In the United States, the cost of laboratory tests may be billed to medical insurance and this billing process may make the process of ordering tests cumbersome for dental providers. For instance, the ordering dental healthcare worker may not be a preferred provider with the medical insurance company. The definition of a preferred provider is a provider who has a contract with the patient’s health insurer or plan to provide services at a discount. For nonpreferred providers, the fees charged for laboratory tests ordered may be costlier and the patient may be responsible for a larger portion of the charges than if the provider had been a preferred provider (in the network). The laboratory charges to the patient in a situation such as this may be prohibitive, and the patient may not proceed with recommended dental treatment secondary to the laboratory costs, or may be upset with the dental team when the bill from the diagnostic laboratory is received. In such cases, it may be more efficient and less costly for a patient with medical insurance to have the necessary laboratory tests ordered by their primary care physician. The dental healthcare worker should communicate with the physician when requesting labs and discuss the planned dental procedure and need for the test. Similarly, the physician will need to update the patient’s medical records with the reasons the tests were ordered and share those results with the dental healthcare worker.


Laboratory results are reported with reference intervals, which are more commonly known as reference ranges. Reference ranges aid in the interpretation of laboratory results. Reference ranges for each test are determined using the demographics of the presumed healthy population from which specimens were obtained and the specific methods and/or instruments used to assay these specimens. The use of population reference ranges is an inexact science, and factors such as age, sex, and ethnicity are generally not considered. Reference ranges are usually defined as the set of values that 95% of the normal population falls within (also known as a 95% confidence interval). Reference ranges can be influenced by many factors.

Laboratory values that are within the reference range do not assure health. Likewise, laboratory values that fall outside of the reference range may not confirm disease or indicate a problem. When multiple tests on an individual are performed, there is an increased likelihood that an abnormal value may be due to chance. Lab values outside of the reference range should be investigated when clinically or statistically significant. Reference range values may differ among laboratories. Laboratories that are accredited by the College of American Pathologists (CAP) are required to establish and/or validate their own reference values at least annually.18 Variance in reference ranges from laboratory to laboratory may occur for a variety of reasons, such as the equipment used in the analysis, reagents used, site humidity, temperature differences, and other conditions.


Formed elements of the blood arise from a common pluripotent hematopoietic progenitor cell, which matures and differentiates to form the various cellular elements. These cells include erythrocytes (red blood cells, RBCs), leukocytes (white blood cells, WBCs), and platelets.

Of these elements, RBCs are the most numerous. RBCs are required for the delivery of oxygen to tissues and use the iron‐containing protein hemoglobin to transport oxygen and carbon dioxide. RBCs, unlike WBCs, have no nuclei (anucleate).

Analysis of RBCs includes the volume of packed red cells (hematocrit), concentration of hemoglobin (Hb), and concentration of red cells per unit volume (RBC count; see Table 28‐3). Additionally, three indices are used to describe the quality of the red cells sampled. These indices are the mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), and mean corpuscular hemoglobin concentration (MCHC).

Table 28‐3 Red blood cell count.

Specimen Traditional Reference Interval SI Reference Interval
Female whole blood 3.9–5.5 ×106 μL 3.9–5.5 ×1012/L
Male whole blood 4.6–6.0 ×106 μL 4.6–6.0 ×1012/L

Hemoglobin Concentration

Hb is measured through spectrophotometric methods secondary to its intensely colored nature. Its main function is to serve as an oxygen carrier and it is found in a variety of forms within the blood. These forms include methemoglobin, oxyhemoglobin, carboxyhemoglobin, and other minor components. In order to measure Hb, whole blood is mixed with Drabkin’s solution. Drabkin’s solution contains sodium bicarbonate, potassium ferricyanide, and potassium cyanide, which causes a conversion of the Hb forms to cyanmethemoglobin. The light absorbance of cyanmethemoglobin is then measured using a spectrophotometer at 540 nm and the concentration of Hb is determined in grams per deciliter of blood (g/dL) (see Table 28‐4). Whole blood samples are collected in a purple/lavender‐top tube using EDTA (see Table 28‐2) as the anticoagulant and may be rejected by the laboratory if the sample has hemolyzed or clotted. Hemolysis (hemolyzed samples) is defined as a pathologic breakdown process in blood of the RBCs. Hemolysis is typically accompanied by varying degrees of a red‐colored tinge in serum or plasma once the whole blood specimen has been centrifuged.

As discussed previously, a subfraction of normal Hb is glycosylated hemoglobin or HbA1c, which is formed during the maturation of the RBC. HbA1c is normally about 4.0% of all hemoglobin in adults, but may be elevated to two or three times normal level in individuals with diabetes mellitus. During the maturation process of RBCs, a glucose molecule is attached to the Hb β‐polypeptide chains through a nonenzymatic reaction. The rate of synthesis of HbA1c is dependent on blood glucose levels and therefore monitoring HbA1c levels gives a good indication of average blood glucose levels of the preceding 2–3‐month time period. As such, these values have become useful in evaluating long‐term blood glucose levels in patients with diabetes mellitus (Table 28‐5).

Table 28‐4 Reference range values for hemoglobin (Mass Concentration).

Traditional Reference Interval SI Reference Interval
Male whole blood 13.5–17.5 g/dL 135–175 g/L
Female whole blood 12.0–15.5 g/dL 120–155 g/L

Table 28‐5 Reference range values for glycosylated hemoglobin.

Traditional Reference Interval SI Reference Interval
Whole blood 4.0%–5.6% of total hemoglobin 4.0–5.6 mmol/L

Table 28‐6 Reference ranges for hematocrit.

Specimen Traditional Reference Interval SI Reference Interval
Male whole blood 41%–50% 0.41–0.50
Female whole blood 35%–45% 0.35–0.45


The hematocrit (Hct) may be measured either using an automated counter or manually. Manual measurement entails centrifugation of anticoagulated whole blood and this is generally an accurate measure of red cell status. Generally, the anticoagulant EDTA is used and a sample is drawn into a purple/lavender‐top tube (see Table 28‐2). There are certain inherent technique errors in manual determination, though, that may affect the results. Specifically, manual determination measures red cell concentration and not red cell mass. This factor may be important when dealing with a patient who is hemoconcentrated as a result of shock or volume depletion. In these patients, red blood cell volume may be normal or high, but red cell mass would actually be decreased.

Automated measurement of Hct usually does not depend on centrifugation and the automated values closely parallel the manual values. The automated technique directly calculates the Hct through determining the red cell number and red cell volume. In this technique, red cell number multiplied by red cell volume equals Hct (Table 28‐6).

Hct values are used in calculating the MCV and the MCHC.

As the amount of RBCs contained within the body is volume dependent, there are differing reference ranges for ages and sexes. Specimens may be rejected from the laboratory if they do not contain enough of a sample or if the sample has coagulated or hemolyzed before analysis.

Red Cell Indices

Red cell indices are useful when evaluating patients for anemias. MCV evaluates for the size of the average RBC, MCH evaluates for the weight of Hb, and MCHC evaluates for the amount of Hb present compared to size.

Mean Corpuscular Volume

The average volume of the RBC is measured through MCV (Table 28‐7). This index is used in the classification of anemias. MCV may be calculated using the following formula:

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Nov 28, 2021 | Posted by in General Dentistry | Comments Off on Laboratory Medicine and Diagnostic Pathology
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