Core Area One: Renal

Group A

Analgesic Nephropathy

The Jaffe Method

Enzymatic Assays

Analgesic Nephropathy

• A chronic kidney disease that gradually leads to end-stage renal disease (ESRD) and the need for permanent dialysis or a kidney transplant to restore renal function.

• Its likely cause is the habitual use of compound analgesics such as acetaminophen and aspirin.

• Predominates in women peak incidence at age 50-55 years.

• Symptoms usually develop after a cumulative analgesic dose of 2-3 Kg.

Manifestations of Analgesic Nephropathy

• Acute renal failure: sudden loss of the ability of the kidneys to remove wastes, concentrate urine and conserve electrolytes:

-Urinalysis may be normal.

-Serum Cr may increase.

-Blood Urea Nitrogen (BUN) may increase.

-Serum K+ may increase.

- Metabolic acidosis may develop.

Manifestations of Analgesic Nephropathy

• Chronic Renal failure: Gradual and progressive loss of the ability of the kidneys to excrete wastes, concentrate urine and conserve electrolytes:

-Urinalysis may show protein or other abnormalities.

-Serum Cr progressively increases. -BUN progressively increases. - Serum K+ levels increase. -Metabolic acidosis develops.

Manifestations of Analgesic Nephropathy

• Interstitial Nephritis: Inflammation of the tubules and the spaces between the tubules and the glomeruli:

-Small amounts of protein and sometimes red blood cells.

-WBCs, WBC casts, RBCs may appear in urine.

-Urine pH raised and metabolic acidosis.

-BUN and serum Cr increase.

Manifestations of Analgesic Neohropathy

• Renal papillary necrosis.

• Urinary tract infections.

• Hypertension.

The Jaffe Method: Principle of Operation

• Jaffe reaction:

Creatinine+Picrate----> Complex

• (Alkaline conditions)

• Complex is red-orange, easily detected and quantified.

The Jaffe Method:Positive Interference

• Positive interference in normal subjects of up to 20% caused by non-creatinine chromogens :

-glucose, ascorbate, uric acid etc. that slowly reduce alkaline picrate.

-Acetoacetate, pyruvate and other ketoacids and proteins that form colored complexes with alkaline picrate.

• This leads to the GFR being underestimated. However these interferences are accounted for in normal subjects BUT become significant in diseased individuals with ketoacidosis and cephalosporin treatment.

The Jaffe Method: Negative Interference

• Caused by high bilirubin levels as well as other compounds in the serum of jaundiced patients.The mechanism of action is not fully understood.

• GFR is overestimated as the measured creatinine concentration is reduced.

The Kinetic Jaffe Method

• Rate of color development of creatinine is different to that of non-creatinine chromogens.

• Creatinine can now be seperated from interfering substances as reactions are rate-dependent.

• Positive interference is significantly reduced however it is not eliminated. (This reduces the reference range for this method by 20% cf the standard Jaffe Method which is a problem when comparing results from different labs)

• Negative interference is still significant.

Enzymatic Assays: Principle of Operation

• A series of sequential enzyme mediated steps.

• Initial step: degradation of creatinine.

• Final step: Enzymatic production of a quantifiable product.

• Can be a one step assay.

Enzymatic Assays: Principle of Operation

A Few Examples:1. Cratinine PAP Method:• Creatinine---->Creatine (Creatininase)• Creatine---->Urea+Sarcosine (Creatinase)• Sarcosine---->Formaldehyde+Glycine

+Hydrogen Peroxide (Sarcosine Oxidase)• Hydrogen Peroxide+Indicator---->Red

Benzoquinone-imine dye (A substance that absorbs at a maximum of 510 nm)

Enzymatic Assays: Principle of Operation

1. The Ektachem Method:

• Creatinine---->N-methyldantoin +Ammonia

• (Ammonia is quantified via a reaction with Bromophenol)

Enzymatic Assays: Advantages

• Precise

• Accurate

• Practical

• Easily adaptable

• Use a small sample size

• Rapid

• Show Clinical specificity

Enzymatic Assays: Disadvantages

• Expensive

• Interference

• Variation in reference ranges

• Imprecise at lower serum creatinine concentrations

Jaffe Method Comparison to the Enzymatic Assay Method

1. Relative Accuracy:• Same accuracy in normal individuals• More accurate in diseased individuals allowing: -No no interference from increased

glucose levels -No interference from increased ketone

bodies -No interference from cephalosporins -Reduced interference from increased

bilirubin levels

Jaffe Method comparison to the Enzymatic Assay Method

• (Applies to the PAP method, as other assays differ in level of intrference or type of interfering substance)

Clinical Implications of Discrepancies Between the two Protocols

• The enzyme assay method, due to no or reduced interference, becomes a more diagnostic laboratory tool for the measurement of serum creatinine levels in patients with diabetes mellitus, neonates, and patients receiving cephalosporins.

Conclusion

• The Jaffe Method has both positive and negative interference issues especially problematic in diseased patients suffering from ketoacidodsis, jaundice or undergoing cephalosporin treatment.

• The kinetic Jaffe Method overcomes the positive interference issues but not the negative interference issues and can’t be used in jaundiced patients..

• The Enzyme Assay Method overcomes both the positive and the negative interference issues and is a better diagnostic tool to use in diseased patients.

• Standardization is required for reference ranges of the different assays.