1. Pop- quiz

2. Q1- Identify 1-6 in the given illustration2 1 3 4 5 6

3. Q2-In the given light spectrum:
Region 1 represents…………………..
Region 2 represents…………………..
Region 3 represents…………………..

4. Q3- According to the Beer-Lambert Law, Absorbance is dependent on :
A) Distance traveled by the light through the sample.
B) Extinction coefficient of the sample.
C) Solution concentration.
D) A and C.
E) A, B and C.
The Beer-Lambert law (or Beer's law) is the linear relationship between absorbance and concentration of an absorbing species. The general Beer-Lambert law is usually written as: A = a(lambda) * b * c where A is the measured absorbance, a(lambda) is a wavelength-dependent absorptivity coefficient, b is the path length, and c is the analyte concentration. When working in concentration units of molarity, the Beer-Lambert law is written as: A = epsilon * b * c where epsilon is the wavelength-dependent molar absorptivity coefficient with units of M -1 cm -1 .

5. Collect the Answer Sheets

7. ANSWERS

8. Q1-Identify 1-6 in the given illustration
1) Light source
2) Wavelength Selector
3) Slit
4) Cuvette
5) Light-Detecting Photocells
6) A scale or a meter
Q2-In the given light spectrum:
Region 1 represents: infrared light
Region 2 represents: Visible light
Region 3 represents: Ultraviolet
Q3-A, B and C.

9. Renal function tests

10. Outline the components of renal function test.
By the end of this topic, the student will be able to
Outline the components of renal function test.
Estimate the concentration of creatinine in a serum sample using a colorimetric method, Jaffe´ Reaction.
Interpret the resulting data.
Calculate creatinine clearance.
Identify their clinical use.

11. INTRODUCTION
The kidneys have several vital roles in maintaining health.
Filter waste materials from the blood and expel them from the body as urine.
They help to control the levels of water and various minerals in the body.
They are critical to the production of vitamin D, red blood cells and hormones that regulate blood pressure.

12. Urine analysis Glomerular function
Types of Renal function tests:
It assess the chemical and physical characters of the urine.
Urine analysis
Blood Urea Nitrogen (BUN).
Estimation of serum creatinine.
Creatinine clearance test.
Glomerular function

13. Blood Urea Nitrogen (BUN).
Urea is a nitrogen containing compound formed in the liver as the end product of protein metabolism.
It is eliminated in the urine as a major nitrogen waste product (85%).
It is often used as an index of renal function along with plasma creatinine.
BUN checks for waste products in the blood.
BUN measures the amount of nitrogen in the blood.

14. normal BUN levels fall in the following ranges from 6 to 20 mg/dL
Normal value of serum BUN:
normal BUN levels fall in the following ranges from 6 to 20 mg/dL

15. Blockage of the urinary tract. Low blood flow to the kidneys .
High BUN value may be caused by:
kidney disease .
Blockage of the urinary tract.
Low blood flow to the kidneys .
Many medications.
High-protein diet, tissue damage and bleeding in the gastrointestinal tract.

16. Very low protein diet, malnutrition, or severe liver damage.
Low BUN value may be caused by:
Very low protein diet, malnutrition, or severe liver damage.
Women and children may have lower BUN levels than men

17. Colorimetric estimation of serum creatinine

18. Aim of practical session
The aim of this practical session is to:
Estimate the concentration of creatinine in a serum sample using a colorimetric method, the Jaffé reaction.
Comment on the provided case.
Calculate creatinine clearance.

19. Introduction
Creatinine is a non-protein waste product of creatine phosphate metabolism by skeletal muscle tissue.
Creatinine production is continuous and is proportional to muscle mass.

20. PRINCIPLE
The assay is based on the reaction of creatinine with sodium picrate as described by Jaffé. .

21. Principle of the Jaffé reaction
In the Jaffé reaction, creatinine reacts with alkaline picrate solution to yield a deep orange-red complex

22. Principle of the Jaffé reaction
Intensity of the colour formed is directly proportional to the amount of creatinine present in the sample.

23. For the Standard sample
Procedure
Step 1:Label 3 dry test tubes B
For the Blank T
For the test sample S
For the Standard sample

24. B T S Alkaline picrate 1.0 ml Distilled water 2.0 ml …….. Test
Step 2- Pipette into the labeled test tubes B T S
Alkaline picrate
1.0 ml
Distilled water
2.0 ml
……..
Test
Standard

25. Step 3: Pipette 2 ml of the given Test sample ,Standard and DW into their corresponding tubes
0.9
2 ml

26. Step 4: for all ; Pipette 1ml of the alkaline picrate reagent into the previous 3 test tubes.

27. Step 5: Mix the contents of the tubes

28. let the tubes to stand for 20 minutes at room temperature.

29. Record the absorbance of both test & standard against the blank using the Wavelength : 520 nm.

30. Calculate the test concentration by the following equation:
Concentration of serum creatinine (mg/dL) =
Absorbance of Test
X Concentration of standard
Absorbance of standard

31. Normal value of serum creatinine
Adult males: mg/dl: values are slightly higher in males due to larger muscle mass.
Adult females: mg/dl.
Children: mg/dl

32. Impaired renal function. Chronic nephritis. Urinary tract obstruction.
Increased serum creatinine levels are seen in:
Impaired renal function.
Chronic nephritis.
Urinary tract obstruction.
Muscle diseases such as gigantism, acromegaly, and myasthenia gravis.
Congestive heart failure.
Shock.

33. * Persons with small stature. * Decreased muscle mass.
Decreased serum creatinine levels are seen in:
* Persons with small stature.
* Decreased muscle mass.
* Inadequate dietary protein.
* Muscle atrophy.

34. Determination of Creatinine clearance
Creatinine clearance rate is the volume of blood plasma that is cleared of creatinine per unit time and is a useful measure for the
Glomelular Filteration Rate.

35. Steps:
1-Advise the patient to avoid eating
much protein diet and not to use any
type of diuretic drugs or coffee.
2- Collect the 24 hours urine ,measure
its volume and a blood sample is taken.

36. 3- Estimate the concentration of
creatinine in serum and in the total
urine sample .
4- Calculate creatinine clearance from
the following equation;

37. Clearance is measured as milliliters/minute (ml/min) =
CALCULATION
Clearance is measured as milliliters/minute (ml/min) =
Urine creatinine X 24 hs urine volume in ml
Serum creatinine X 24 X 60

38. Normal value of serum Creatinine clearance
Adult males: to 137 ml/min.
Adult females: 88 to 128 ml/min.
Children: to 140 ml/min.

39. Decreased creatinine clearance levels are seen in:
Acute tubular necrosis.
Congestive heart failure.
Dehydration.
End-stage kidney disease.
Glomerulonephritis.
Kidney failure.
Renal ischemia (too little blood flow to the kidneys).
Renal outflow obstruction (usually must affect both kidneys to reduce the creatinine clearance).
Shock.

40. Microalbumin-Creatinine ratio
The body normally filters out creatinine in the urine at a steady rate
Comparing the ratio of urine albumin with creatinine in the same urine specimen helps determine if the body is excreting albumin at an increased rate.
If this is happening, it may be the result of kidney disease.

41. Evaluating patients with Diabetes for kidney disease
The presence of small amounts of albumin in the urine is called microalbuminuria and is associated with early-stage kidney disease.
Once there are larger amounts of albumin in the urine it is called macroalbuminuria and it could indicate more severe kidney disease.
Evaluating patients with Diabetes for kidney disease

42. Steps:
1-measuring the amount of albumin in the urine
2-measuring the amount of creatinine in the urine
3-The amount of urine albumin is compared with creatinine.

43. Microalbumn urine in mg/dL Creatinine urine in mg/dL
Calculation:
Microalbumn urine in mg/dL
Creatinine urine in mg/dL
Dividing the albumin by creatinine gives: mg/mg
Multiply that by 1,000 to get
mg albumin/g creatinine

44. normal less than 30 mg microalbumin/g creatinine
Normal value of microalbumin/creatinine:
normal less than 30 mg microalbumin/g creatinine

45. Lab activity
Calculate serum creatinine concentration in one of the two provided adult male serum samples.
(Normal serum creatinine in Adult males: mg/dl)
2.Comment on your result.

46. Concentration of serum creatinine (mg/dL) =
Absorbance of Test
X Concentration of standard
Absorbance of standard

47. Thank you