Hemolysis is the rupture of red blood cells, releasing their intracellular contents, such as hemoglobin, into the plasma or serum. When this occurs, the sample liquid changes from a clear, straw-yellow color to various shades of pink or red, depending on the extent of cell damage. Preventing this damage is important because hemolyzed samples introduce pre-analytical error, leading to inaccurate test results. Tests for potassium, lactate dehydrogenase (LDH), and bilirubin are highly susceptible to interference from the released cellular contents, often requiring the sample to be recollected and delaying patient diagnosis and treatment.
Pre-Collection Preparation and Equipment Selection
Preventing red blood cell damage begins before the venipuncture. Selecting the correct needle gauge is crucial, as using one smaller than 23 gauge increases shear stress on cells passing through the narrow opening. For routine venipuncture, a needle gauge in the range of 20 to 22 is recommended to balance patient comfort with optimal blood flow. Conversely, using an overly large needle can cause trauma to the vein wall or increase flow turbulence, which also contributes to cell rupture.
The tourniquet should be applied for the shortest duration possible, ideally no longer than 60 seconds. Prolonged application causes blood stagnation and hemoconcentration, artificially elevating the concentration of larger blood components and increasing the risk of mechanical damage to the cells during the draw.
Residual alcohol on the skin can enter the sample and cause chemical hemolysis. Healthcare professionals must ensure the cleaning agent is completely air-dried before the needle is inserted. All collection tubes should be checked for their expiration date and filled to the appropriate volume, as underfilling tubes with anticoagulants can lead to an excessive concentration of the additive, which promotes cell breakdown.
Optimizing Needle Insertion and Angle
The physical act of needle insertion requires a technique that minimizes trauma. The insertion should be smooth, swift, and deliberate, avoiding slow movements that can scrape the vessel lining. A clean puncture is less likely to damage the delicate cell membranes, which are highly sensitive to mechanical stress.
The ideal angle for needle insertion is between 15 and 30 degrees. This angle ensures the bevel enters the vein completely and remains fully seated within the vessel lumen. If the angle is too shallow, the bevel may partially rest against the vein wall, causing friction and subsequent cell damage as blood is drawn.
Excessive movement, repositioning, or “fishing” for the vein significantly increases the risk of mechanical hemolysis. If the blood flow becomes sluggish or the initial attempt is unsuccessful, discontinue the draw, remove the needle, and attempt a new puncture at a different site using fresh equipment. This prevents the turbulence and shear stress that occurs when the needle is partially occluded or adjusted against the vessel wall.
Managing Flow Rate and Vacuum Pressure
During collection, the force applied to the blood cells is a primary determinant of hemolysis. If a syringe is used, the healthcare provider must pull back on the plunger gently and slowly. Rapid aspiration creates excessive negative pressure, forcing the red blood cells through the needle at high velocity, which causes damage due to shear stress.
When using an evacuated tube system, the vacuum pressure is predetermined. It is important to match the tube size to the vein’s capacity, as the strong vacuum in a standard tube may collapse a small or fragile vein. Vein collapse leads to turbulent blood flow and increased friction against the needle opening, both high-risk factors for cell rupture.
When drawing blood from an existing intravenous (IV) line or catheter, the initial aspiration must be done very gently, as forceful aspiration through these narrow, high-turbulence systems is a major cause of hemolysis due to high shear forces. Obtaining the sample via direct venipuncture is preferred, as straight needle draws significantly reduce hemolysis rates compared to draws from IV starts.
Post-Collection Handling and Transport
Tubes containing additives, such as anticoagulants or clot activators, must be mixed immediately to ensure the substance is evenly distributed. This mixing must be performed by gentle inversion, typically 5 to 10 times depending on the tube type, rather than vigorous shaking. Vigorous shaking is a common cause of mechanical hemolysis, as the rapid agitation and impact of the blood against the tube walls destroys the fragile red cells.
If the blood was collected using a syringe, it must be transferred correctly. The needle should be removed from the syringe, and a specialized transfer device should be used, or the blood should be gently dispensed down the side of the tube. Forcing the blood through the syringe needle into the tube, especially by pushing hard on the plunger, creates a shearing effect that ruptures the red blood cells.
During transport and storage, samples are vulnerable to temperature extremes and physical agitation. Samples exposed to excessive heat or cold can experience cell breakdown. Samples should be transported gently, avoiding physical jarring or violent oscillations that can cause impact rupture. If the sample requires centrifugation, the equipment must be properly balanced and operated at the correct speed and time, as excessive centrifugal force can also induce cell damage.