lipemia. Finally, the plasma sample can be used to assess plasma protein concentration with a refractometer. Normal protein concentrations range from 5.5–7.5 g/dl in dogs and 6.5–8.5 g/dl in cats. Refractometric protein readings can be falsely elevated in samples that are lipemic, hemolyzed, or when there are high concentrations of glucose, urea, sodium, or chloride in the sample (Evans 2011). Complimentary analysis of the PCV and plasma protein can provide useful information to narrow the list of differential diagnoses:
Low PCV, Normal protein: anemia
Low PCV, Low protein: chronic blood loss
High PCV, Normal protein: increased red blood cell production, hemorrhagic gastroenteritis, endocrinopathy
Normal PCV, Decreased protein: protein‐losing disease, acute blood loss, liver failure
Normal PCV, High protein: dehydration, increased immune response.
Urea is a waste product of dietary protein digestion formed in the hepatocytes and excreted through the kidneys. As such, assessment of the BUN concentration can be a useful indicator of decreased glomerular filtration and, when interpreted in light of the animal's hydration status and urine specific gravity, a crude screening test of renal function. Outside of a concentration reported from a biochemical analyzer, BUN levels are commonly assessed through the application of a whole blood sample on semi‐quantitative colorimetric reagent test strips. Such strips have been correlated with serum urea concentrations as measured with an automated analyzer in both dogs and cats. Dogs with BUN concentration estimates >15 mg/dl and cats with estimates >50 mg/dl were accurately categorized as azotemic in 98% of samples evaluated (Berent et al. 2005).
A final point‐of‐care quick assessment test is the measurement of blood glucose concentration. Glucose is the primary cellular energy source and is obtained from dietary carbohydrates, the breakdown of glycogen in the liver (glycogenolysis) and the synthesis of glucose from amino acids and fats (gluconeogenesis). Alterations in blood glucose concentration can indicate dysfunction in a wide variety of organ systems including the renal, hepatic, and endocrine systems. In addition, sepsis and congenital metabolic diseases are important causes of hypoglycemia while analysis immediately or shortly after a meal or a stressful event (particularly in cats) are noteworthy explanations for hyperglycemia (Duncan 1998). Normal fasting blood glucose concentrations range from 60 to 125 mg/dl in dogs and 70 to 150 mg/dl in cats.
Care must be taken when obtaining a blood sample for the measurement of glucose concentration. Acute stress, such as that due to aggressive handling or restraint, and a recent (<12 hours) meal are likely to result in falsely elevated measurements. Additionally, blood glucose concentration decreases ~10% per hour at room temperature, so samples should be processed within 30 minutes or the serum should be separated and refrigerated until testing (Evans 2011).
Glucose concentrations in whole blood can be measured by using colorimetric reagent strips or portable blood glucose meters. Reagent strips are not reliable for the detection of hypoglycemia, and the presence of anemia may result in falsely elevated concentrations. Excessive washing and incomplete coverage of the reagent pad can also result in erroneous results. Elevated blood glucose concentrations above the renal threshold can also be detected in urine samples through the use of urine chemical reagent test strips (Evans 2011). Portable blood glucose meters can provide measurements that are comparable to those obtained with reagent strips and biochemical analyzers; however, there is substantial variation between models (Cohen et al. 2009, 2000). In general, such meters tend to underestimate blood glucose concentrations and have greater divergence from reference measurements when samples are hyperglycemic. Meters that are designed and calibrated specifically for use in companion animals are preferred and thought to be less likely to report falsely decreased blood glucose concentrations (Kang et al. 2016). As with reagent strips, anemia can result in an overestimation of the blood glucose concentration. Despite these pitfalls, imprecise results from portable blood glucose meters are not likely to lead to alterations in clinical decision‐making (Cohn et al. 2000; Wess and Reusch 2000a, b).
4.3.3.2 Additional Secondary Tests
Several additional diagnostic tests can be conducted in‐house depending on both facility and veterinary resources. Common examples of such tests include diagnostic imaging and necropsy. Many shelters have the capability to perform diagnostic imaging in the form of radiographs (X‐rays) and/or ultrasounds. Such tools can be rapid and useful methods of establishing diagnoses and identifying emergent conditions. In order to ensure that images are of diagnostic quality and, in the case of radiographs, that staff and patient exposure to radiation are minimized, these tools should only be used by appropriately trained personnel; in some jurisdictions, specific licensing requirements may restrict which staff members can take radiographs. Diagnostic images require interpretation by a veterinarian.
Finally, in the event of an unexplained patient death, a gross necropsy should be conducted. Frequently overlooked as a diagnostic tool, a necropsy can help establish a diagnosis and alert the practitioner to any conditions that may impact both the human and the remaining animal population within the shelter. The collection of tissue samples for histopathologic examination and ancillary diagnostic testing (bacteriology, virology, cytology) should also be considered based on gross necropsy findings. Samples can always be collected and held for submission pending initial results, but failure to collect appropriate samples at the time of necropsy may preclude obtaining a definitive diagnosis and significantly limit the information that can be obtained from the necropsy. See Chapter 5 for more information on this topic.
4.3.4 Diagnostic Laboratory Tests
4.3.4.1 Serology
Serology is the measurement of antigen–antibody interactions for diagnostic purposes and is commonly used for the diagnosis of infectious disease as well as risk management during a disease outbreak. In some cases, serology is used to determine whether an animal has been previously exposed to an infectious agent, vaccinated, or actively infected. Primary binding tests are those that directly measure the binding of antigen to antibody, usually in a qualitative form (i.e. positive or negative). Examples of these include ELISAs and lateral flow assays, immunofluorescent antibody (IFA) tests, Western blotting, and immunohistochemistry. Secondary binding tests measure the results of antigen–antibody interaction in vitro and typically provide a quantitative result in the form of a titer; examples include hemagglutination inhibition (HI) and complement fixation (CF). Tertiary testing methods, such as serum neutralization, measure the protective effect of antibodies in live organisms and also provide results in the form of titers. A final category of serologic tests is the molecular assay that detects nucleic acids. These tests, which can be both qualitative and quantitative, include PCR, reverse transcriptase PCR, and real‐time PCR (Tizard 2013).
4.3.4.1.1 Primary Binding Tests (Note that ELISAs including lateral flow assays are common primary binding tests; these have been discussed earlier in the text.)
IFA tests can be either direct or indirect. Direct fluorescent antibody tests can detect the presence of specific antigens within a tissue sample. With IFA, antibody labeled with a fluorescent marker is applied to a tissue sample that has been treated with antibody specific to the antigen of interest. The fluorescent marker binds to the antibody, revealing the presence of antigen when examined under a dark‐field microscope with an ultraviolet light source. This method is commonly used for the detection of rabies virus in brain tissue and FeLV in white blood cells. Indirect fluorescent antibody tests can be used to detect the presence of either tissue antigen or, more commonly, serum antibody. This technique is similar to direct fluorescent antibody testing, however, the sample to be tested is treated with antibodies against the molecule of interest prior to fluorescent labeling. This allows for amplification of the fluorescent signaling, determination of specific classes of antibody in the sample (i.e. IgG, IgM, IgA, etc.), and a quantitative estimation of the amount of antibody in the sample (Tizard 2013). Indirect fluorescent antibody tests are commonly used for the detection of brucellosis