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The Clinical Chemistry Lab

The Clinical Chemistry Lab. Vicki S. Freeman, Ph.D. Objectives. Upon completion of lecture, discussion, case studies and laboratory, the student will be able to: Quality Control and CLIA Regulations Explain the importance of QC in the lab

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The Clinical Chemistry Lab

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  1. The Clinical Chemistry Lab Vicki S. Freeman, Ph.D.

  2. Objectives Upon completion of lecture, discussion, case studies and laboratory, the student will be able to: • Quality Control and CLIA Regulations • Explain the importance of QC in the lab • Define sensitivity, specificity, shift, trend, precision , accuracy and reliability • Describe the 6 aspects of quality control • Classify procedures according to the CLIA regulations • List the Quality Assurance requirements mandated in the CLIA regulations • Carbohydrates • Differentiate types of diabetes by clinical symptoms and laboratory data • Type 1 • Type 2 • Gestational diabetes • Impaired glucose tolerance

  3. Objectives (Con’t) Upon completion of lecture, discussion, case studies and laboratory, the student will be able to: • Carbohydrates (Continued) • Relate expected laboratory results and clinical symptoms to the following metabolic complications of diabetes: • Ketoacidosis • Hyperosmolar coma • Describe the used of hemoglobin A1C and microalbumin in the long term monitoring of diabetes • Lipids and Cardiac Risk • Discuss cholesterol, LDL cholesterol, HDL cholesterol, lipoproteins and triglycerides as predictors of cardiovascular risk • Calculate a LDL cholesterol, given total cholesterol, triglyceride and HDL results

  4. Quality Control • Purpose of QC is to • assure the reliability of patient data obtained from a procedure • monitor variables that can alter data • Patient data is unknown information and “known” samples must be run concurrently

  5. Terms • Precision • the reproducibility of the result • Accuracy • the closeness of the measured result to the true value • Reliability • the ability to maintain both precision and accuracy

  6. Quality Control • Precision • Accurate • Reliability

  7. Shift • 6 or more consecutive values distributed above or below the mean • Trend • A continual increase or decrease in 6 or more consecutive values

  8. Quality Control • Shift • Trend • Outliers

  9. 6 Aspects of Quality Control • Sample collection • Method of analysis • Proper control material • QC monitoring • Instrument maintenance • Documentation

  10. CLIA ‘88 • Waived tests • Moderate complexity tests • High complexity tests • Provider-performed microscopy

  11. UA dipstick Spun hematocrits Hemoglobin Sedimentation rate Fecal occult blood UA pregnancy test Ovulation tests Single analyte instrument Hemacue Glucose Total cholesterol Waived Test Category

  12. Wet mounts vaginal cervical skin KOH preparations Pinworm exams Fern tests UA sediment exam Postcoital direct vaginal and cervical mucus Nasal granulocytes Fecal leukocytes Qualitative semen analysis Provider-performed Microscopic

  13. Hemoglobin Hematocrit Dipstick UA Occult Blood Strep A (ag) UA pregnancy Glucose Cholesterol Triglycerides BUN Uric Acid Cholesterol HDL Cholesterol Common Tests in POLs

  14. Quality Assurance Vicki S. Freeman, Ph.D.

  15. Quality Assurance Program • Written laboratory procedure manual • Specimen collection and identification • Methodologies • Reference Ranges • Quality control • Preventive maintenance • Record keeping

  16. Carbohydrates Lipids Enzymes Cardiac Function Liver Function Renal Function Electrolytes Parathyroid Function/ Calcium Metabolism Acid/Base Balance Pancreatic Function Prostate Profile Groups

  17. Carbohydrates Hyper and Hypoglycemia

  18. Classes of Hyperglycemia • Diabetes Mellitus • Impaired Glucose Tolerance • Gestational Diabetes

  19. Diabetes • A syndrome characterized by inappropriate hyperglycemia with chronic microvascular complications. • Upper limit of 100 mg/dl on the FPG as the upper limit of normal blood glucose

  20. Diabetes Mellitus • Types • Type 1 (Type I) -Insulin Dependent Diabetes Mellitus • IDDM • Type 2 (Type II) -Non Insulin Dependent Diabetes Mellitus • NIIDM • Other Types - Secondary

  21. TYPE 1 TYPE2 Destruction of the B-cells Resistance to Insulin Absolute Deficiency of Insulin Relative Deficiency of Insulin

  22. Type 1 (Type I) - IDDM • Characteristics • Abrupt Onset • Insulin Dependent • Ketosis-prone • Genetic related - HLA Dw4 related • Islet Cell Antibodies • 10 - 20 % of all diabetes

  23. Type 2 (Type II) - NIDDM • Characteristics • Little or no symptoms • Does not exhibit the characteristics of IDDM • Have high basal insulin levels - develop insulin resistance • Decreased # of insulin receptors in insulin-sensitive tissues • Subclasses • Non-obese • Obese - 60 - 90 % of all diabetics • Further divided by the type of treatment the patient receives (Insulin, oral hypoglycemic, diet)

  24. Other Types - Secondary • Destroyed pancreas • Pituitary Hyperfunction • Cushings Disease

  25. Diagnosis of Diabetes Mellitus • Classic symptoms & a casual plasma glucose concentration>200 mg/dl • Fasting venous plasma glucose concentrations >126 mg/dl • 2 hour post prandial >200 mg/dl • Any of the 3 criteria must be confirmed on a subsequent day by any of the 3 methods.

  26. Diagnosis of Diabetes Mellitus • Classic symptoms of diabetes and hyperglycemia • Laboratory Tests • Preferred - Fasting venous plasma glucose • > 126 mg/dl on more than one occasion • Impaired fasting plasma glucose100 mg/dl - 126 mg/dl • Casual plasma glucose concentration>200 mg/dl • Not recommended - Screening test - 2 hour post prandial • OGTT value of >200 mg/dl in 2 hr sample • Normal <140 mg/dl • Impaired GTT 140 - 199 mg/dl • Abnormal >200 mg/dl • Any of the 3 criteria must be confirmed on a subsequent day by any of the 3 methods

  27. Glucose Curves

  28. Gestational Diabetes Diagnosed by any two of the following: • a fasting plasma glucose of more than 105 mg/dl • a 1-hour glucose level of more than 190 mg/dl • a 2-hour glucose level of more than 165 mg/dl • or a 3-hour glucose level of more than 145 mg/dl

  29. Specimen • Overnight fast • New evidence of diurnal variation with FPG higher in AM • Plasma • NaFloride • if cannot be separated from cells within 60 minutes • Anticoagulated (oxalate) • Whole blood values ~11% lower than plasma • Heparinized plasma values 5% lower than serum • Capillary vs venous blood • Fasting state ~= (2 mg/dL difference) • After a glucose load, capillary values ~20-25% higher

  30. Monitoring Therapy • Day - to - day control • Self Monitoring Blood Glucose (SMBG) • Long term • Hemoglobin A1C • Shows a direct relationship with the glucose level over the preceding 2-3 months • Microalbumin • Monitors Monitors kidney function • Urine glucose - obsolete • Urine ketones - fat breakdown products

  31. Glycosylated (glycated) Hemoglobin (GHb or HgbA1c) • ADA Guidelines - Glycosylated hemoglobin • Glucose attaches to tissues and proteins, including hemoglobin • Measures % of hgb that has been modified by glucose • Shows a direct relationship with the glucose level over the preceding 2-3 months • A valuable tool for monitoring glycemia, • Normal levels range from 3%-6% • Should be maintained at <7% (some sources say 6%) • Re-evaluate treatment regimen if GHb >8% • Should be measured at less than 2 x/year (if diabetic is well controlled; otherwise, every 3 months)

  32. Approximate correlation between A1C level and mean plasma glucose levels

  33. Microalbumin • Diabetes is leading cause of end-stage real disease • Microalbumin- Monitors kidney function • Also a marker of increased risk of cardiovascular morbidity and mortality • Annual testing is recommended • Microalbuminuria defined as excretion of • 30-300 mg of albumin/24hrs or • 20-200 g/min or 30-300 g/mg creatinine • On 2 of 3 urine collections

  34. Acute Complications • Ketoacidosis due to lack of insulin/stress, can result in death (assoc. with Type I) • B HCO3-, B pH, A glucose • Hypoglycemia - with too much insulin - results in coma • Hyperosmolar coma (assoc. with Type II) • A blood osmolarity • A glucose

  35. Chronic Complications • Correct Management of diet and insulin, • Avoid further complications of the disease • Retinopathy - blindness 50% after 10 years • Nephropathy - Renal disease • proteinuria, increased BUN and creatinine • Neuropathy - • poor sensation, ulceration of skin, may lead to amputation of limbs • Accelerated macrovascular disease - CAD, CVA

  36. Hypoglycemia • Decreased fasting glucose <50 mg/dl • Fasting • Caused by pituitary/adrenal insufficiency, pancreatic islet cell hyperplasia, islet cell tumors • Other causes may be from drugs, alcohol, insulin • Triad of characteristics (Whipple's Triad) • Hypoglycemic symptoms • Simultaneous demonstration of decreased plasma glucose • Relief of symptoms with glucose administration • Postprandial (or reactive) • Seen after gastric surgery - food is absorbed too fast • Idiopathic - following extreme stress (catecholamines) • Spontaneous recovery

  37. Diagnosis of Hypoglycemia • Diagnosed by looking for the cause • Thrust of the clinical evaluation is to rule out insulinomas. • Hypoglycemia in insulinomas are related to excessive and inappropriate production of insulin - insulin levels are important in making the diagnosis

  38. Diabetes Case Study 1 A 40-year-old African American woman (nonpregnant) has symptoms suggestive of diabetes. On two occasions, the fasting plasma glucose (FPG) is 130 mg/dL and 135 mg/dL. What is the next diagnostic or therapeutic step?

  39. Diabetes Case Study 2 A 35-year-old Caucasian female (nonpregnant) has FPG concentrations on two occasions of 120 mg/dL and 124 mg/dL without symptoms suggestive of diabetes. • How would this patient would be classified?

  40. Diabetes Case Study 3 • A 72 year old male presents with numbness in the legs and frequent urination. A 4 hour glucose tolerance is ordered. The results are: FPG 160 mg/dL 2 hr 220 mg/dL 1/2 hr 205 mg/dL 3 hr 195 mg/dL 1 hr 260 mg/dL 4 hr 165 mg/dL • A follow up Hgb A1c was ordered. • Does this gentleman have diabetes?

  41. Lipids and Cardiac Risk

  42. Cholesterol Synthesis Genomic Medicine: Cardiovascular Disease New England Journal of Medicine Volume 349(1) 3 July 2003  pp 60-72

  43. LipidProfile and Cardiac Risk • Cardiac Risk Factors • Cholesterol, total • Triglycerides • HDL cholesterol • LDL cholesterol (calculated vs direct) • NCEP Guidelines ATPIII • Fasting sample now required for: • Total cholesterol • HDL-C • LDL-C • Triglycerides

  44. Cholesterol • Dependent on many factors • genetics, age, sex, diet, physical activity, hormones • American Heart Association < 200 mg/dl • Measurement • Enzyme method most common

  45. LDL-C • The villain - directly correlated with CHD • Carries cholesterol from its site of origin into the blood vessels • Optimal <100 mg/dL • Near or above optimal 100-129 mg/dL • Borderline high 130-159 mg/dL • High 160 – 189 mg/dL • Very high > 190 mg/dL • Calculation • LDL = Total Cholesterol - (HDL + Triglycerides) 5 Triglycerides > 400 mg/dL causes problem in calculation • Direct measurement (new)

  46. HDL-Cholesterol • The hero - inversely correlated with CHD • transfers cholesterol from cells back to the liver • New!! <45 male; <55 female • Factors which increase HDL • estrogen (women), exercise, alcohol, blood pressure medicine • Factors which decrease HDL include: • progesterone, obesity, smoking, triglycerides and diabetes • Measurement • HDL Precipitation method

  47. Non-HDL-C • Other lipid compounds including • Lp(a), VLDL remnant are significant in individuals with increased triglycerides • Non HDL-C • Triglycerides <200 mg/dL and LDL-C<100 mg/dL • Should then look at non-HDL-C • Total cholesterol – HDL

  48. Triglycerides • Role as a risk factor remains unsettled • Considered an independent risk factor • Definite + association between triglycerides and CHD. >150 mg/dL = risk • While high levels may not cause atherosclerosis, they may indirectly accelerate atherogenesis by influencing the concentration and composition of other lipoproteins • Measurement - fasting > 12 hoursrequired

  49. Other lipid measurements • Lp(a) • similar in structure to LDL • a unique protein apo(a) linked to apolipoprotein B-100 - has a structure similar to plasminogen • directly correlated with CHD - not affected by lifestyle factors such as diet, exercise or smoking • levels >30 mg/dl • Apolipoprotein A • Associated with HDL • Apolipoprotein B-100 • Associated with LDL

  50. Major Risk Factors (Exclusive of LDL Cholesterol) That Modify LDL Goals* • Cigarette smoking • Hypertension (blood pressure > 140/90 mm Hg • or on antihypertension medicine • Low HDL cholesterol (<40 mg/dL) • Family history of premature CHD • males first degree relative <55 years • Female first degree relative < 65 years • Age • Men > 45 years • Women > 55 years • Diabetes is regarded as a CHD risk equivalent • HDL cholesterol > 60 mg/dL counts as a “negative risk factor • Its presence removes 1 risk factor from the total count

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