Interference with alpha-antitrypsin studues in stored serum by presumed bacterial proteases

Area of Science:

• Biochemistry
• Microbiology
• Clinical Chemistry

Background:

• Gram-negative bacterial contamination can compromise serum integrity.
• Spoiled serum impacts measurements of trypsin inhibitory capacity (STIC) and antitrypsin phenotyping.
• Fibrinolytic activity increases in contaminated serum, while antitrypsin levels remain constant by radial immunodiffusion.

Purpose of the Study:

• To investigate the impact of bacterial contamination on serum assays.
• To identify bacterial species capable of spoiling serum for STIC and antitrypsin phenotyping.
• To understand the mechanism of serum degradation by bacterial enzymes.

Main Methods:

• Incubation of human serum with bacterial cultures (Enterobacter, Klebsiella, Bacillus subtilis, Pseudomonas).
• Measurement of trypsin inhibitory capacity (STIC).
• Antitrypsin concentration determination using radial immunodiffusion.
• Assessment of fibrinolytic activity.
• Enzyme activity assays (esterase activity, plasmin activation).

Main Results:

• Bacterial contamination, particularly by Enterobacter, Klebsiella, Bacillus subtilis, and Pseudomonas, leads to serum spoilage.
• Spoiled serum exhibits increased fibrinolytic activity.
• Bacterial enzyme production is enhanced in the presence of human serum.
• The enzyme lacks esterase activity and does not affect STIC when serum plasmin is activated by streptokinase, suggesting a bacterial origin.

Conclusions:

• Bacterial contamination poses a significant risk to the accuracy of STIC and antitrypsin phenotyping assays.
• Preventive measures, including aseptic technique and proper sample handling (rapid freezing, frozen storage/transport), are essential.
• The identified bacterial enzyme contributes to serum degradation, impacting diagnostic reliability.