A “fresh” urine sample is a specimen whose chemical and physical composition accurately reflects the body’s state at the moment of collection. Urine is a complex biological solution containing water, salts, waste products, and cellular elements, and its stability begins to change almost immediately after it leaves the body. Since urine is an ideal culture medium, proper handling is necessary to ensure the integrity of its components for laboratory analysis. Following guidelines for collection and storage is a prerequisite for generating reliable diagnostic information.
Stability Limits at Room Temperature
When a urine sample is left at ambient temperature, its stability is severely limited, typically to a maximum of one to two hours before degradation begins. This short window is due to rapid bacterial proliferation, the most common reason for sample rejection by laboratories. Bacteria naturally present in the urethra or on the skin multiply quickly, dramatically increasing their concentration. This bacterial overgrowth can lead to a false diagnosis of a urinary tract infection if the sample is intended for culture.
Chemical changes also occur quickly without temperature control, primarily driven by multiplying bacteria. Certain bacteria possess an enzyme called urease, which breaks down the urea naturally found in urine into ammonia. This conversion causes the sample’s acidity (pH) to shift rapidly toward alkalinity. Additionally, cellular components and casts, which are structures formed in the kidney tubules, are fragile and begin to break down (lyse) if the urine is not promptly analyzed.
Extending Viability Through Refrigeration and Freezing
Refrigeration is the standard method for extending the viability of a urine sample for short-term storage. Storing the sample at approximately 39°F (4°C) significantly slows the metabolic activity of bacteria and stabilizes many chemical analytes. This allows a urine sample to remain suitable for standard urinalysis for up to 24 hours, though some protocols allow up to 48 hours for certain tests.
While refrigeration does not halt degradation entirely, it merely delays it by inhibiting bacterial growth and slowing the breakdown of cellular components. Even under refrigeration, some cellular elements, like casts, may still gradually break down over the 24-hour period. Because of this continuous, slower degradation, laboratories prefer to analyze the sample as soon as possible after collection.
Freezing is generally reserved for long-term preservation, often for specific chemical analyses or research purposes, and requires a temperature of approximately -4°F (-20°C) or colder. It is highly effective at stabilizing most chemical compounds and metabolites for extended periods, sometimes for several months. However, the freezing process itself can cause significant damage to the physical elements in the urine, such as red blood cells, white blood cells, and epithelial casts.
The formation of ice crystals during freezing physically ruptures the delicate membranes of these cellular elements, causing them to lyse. Consequently, a thawed sample is typically unsuitable for microscopic examination, which relies on intact cells and casts for accurate diagnosis. For any extended storage method, the sample must be collected in a sterile, tightly sealed container to prevent contamination and evaporation, which would otherwise alter the sample’s concentration.
How Sample Degradation Affects Diagnostic Results
Exceeding the recommended time limits for storage, particularly at room temperature, introduces specific changes that can lead to erroneous diagnostic results. One major consequence of delayed analysis is the shift in urine pH toward alkalinity due to the bacterial conversion of urea to ammonia. This increased alkalinity can cause the precipitation of crystals that were not present in the original fresh sample, potentially confusing the diagnosis of kidney or bladder stones.
Furthermore, the alkaline environment can interfere with chemical strip tests, sometimes causing a false-positive reading for protein, which could mistakenly suggest kidney damage. Delayed processing for a urine culture is a common source of error, as bacterial counts increase exponentially. A sample with a low, non-infectious level of bacteria at collection can appear to have a high, clinically significant colony count hours later, leading to a false diagnosis of a urinary tract infection.
The breakdown of formed elements within the urine also results in false-negative findings. Red blood cells and white blood cells, which indicate bleeding or infection, lyse in older samples, reducing their measurable count and masking an underlying condition. Similarly, the fragile protein casts that form in the kidney tubules break apart, causing a false negative result for kidney diseases. Analytes like glucose and ketones degrade over time; bacteria consume glucose, leading to a falsely low reading that could inaccurately suggest a person with diabetes has their condition under control.