ROUTINE URINALYSISI. Macroscopic Examination:Mix the specimen thoroughly and determine the following:A. : COLORLESS (Water Clear): STRAW: YELLOW: DARK YELLOW: LIGHT AMBER: AMBER: DARK AMBER: RED: OTHERS (Specify)B. TRANSPARENCY (Appearance): Clear: Cloudy: SLIGHTLY TURBID: TURBIDC. REACTION (pH): Use Strip testD. PROTEIN (pH): Use strip test. However, if the urine is highly colored such that comparison of the result with the color chart is difficuly, use Heat and Acetic Acid Test. Indicate in the logbook that such method was use for the said determination. E. Sugar (Glucose): Use strip test. However, if the urine is highly colored such that comparison of the result with the color chart is difficult, use the Clinitest. Indicate in the logbook that such method was use for the said determination.F. Specific Gravity: Use the strip testII. Microscopic ExaminationNote: the urine specimen must be examined while fresh, since cells and cast begin to lyse within 1-3 hours.Procedure:1. Mix the specimen well. Pour approximately 10 mL of urine into a test tube. Centrifuge at 2000 rpm for 5 mins. Remove the supernate by decantation. While still inverted, press unto 2 thickness of gauze. Sufficient urine remains in the test tube to suspend the sediment.2. Place a drop of resuspend sediment on one area of a slide and coverslip, avoiding bubbles. Too much fluid will cause the coverslip to float.3. Examine with both low and high power fields. Examine with low power and subdued light. The fine focus should be varied continuously while scanning. Systematically progress around all slides of the coverslip.4. Average the contents of 10 representative fields and report the results. MANNER OF REPORTING:1. Leukocytes (WBC): Count and Record the number of pus cells per HPO. Clumps of pus cells should be reported as such in the same manner.2. Erythrocytes (RBC): Count and Record the number of RBC per HPO,3. Casts and Cylindroids: Count and Record the number of each type per LPO.4. Epithelial Cells: Record the type and amount per hpf.Report as follows:Occasional - less than 5/hpfFew - 5-20/hpfModerate - 20-50/hpfMany - Greater than 50, less than packed/hpf. Loaded - Packed field/ hpf5. Crystals: Record the type and amount per LPO.Report as follows:Occasional - less than 1/lpfFew - 1-5/lpfModerate - 5-10/lpfMany - 10-30/lpfLoaded - greater than 30/lpf6. Miscellaneous Urine Sediment (Bacteria, Yeast, and Spermatozoa)7. Mucus Threads: Report as in item (5)8. Parasites: Report its presence when found in the urine sediment.Trichomonas species- The Trichomonas are seen as pearly shaped trophozoites with undulating membrance and are very motile.Schistosoma haematobium ova- Found in the urine as flask like in shape and taper considerably at one extremity the other being rounded.Echinococci- Occasionally found in the urine. The characteristic parts of the echinococci found in the urine are the hooklets as well as portion of the membrane. Scolices may also be found. These scolices are small and round are supplied with a wreath of hooklets. When echinococci are found in the urine, there is also evidence of hemorrhage or ulceration or both.Red cells are numerous, together with epithelial and connective tissue shreds as well as pus cells. Filaria- appears in the urine at times, causing severe hematouria or chyluria is found, an examination of filarial, it is advisable to take the first urine voided in the morning. Ascaris lumbricoides- Maybe found in the urine.Enterobius vermicularis- Maybe found in the urine.

24 Hours FOR TOTAL PROTEINMethod: Biuret Method Test Principle: Protein is precipitated from the sample by tricholoroacetic acid (TCA) and the determined by the Biuret Method: Normal Value: 25-70mg per 24 hours / 0.025 0.070 gm per 24 hours.Sample: 24 hours Urine Stability of the Sample: It is advisable to spike 24hour urine with 5 mL of 10% thymol in isopropanol solution. The following stability apply:24 hour Urine: Four Days at 15-20 deg CReagents/ Preparation and Stability of Solution:Bottle 1- Biuret Reagent Dilute contents of one bottle with 400mL redistilled water. Rinse thoroughly. Stable for one year at +15-25 deg CBottle 2- Reagent for blank.Do not use solution.Bottle 3- Standard ProteinUse solution undiluted.Stable up to the expiry date specified when stored at +15-25 deg CSample Preparation: Determine the concentration range of protein in the urine specimen using the Test Strips e.g; Multisticks: Protein Concentration: Diltuion: Dilution Factor: < 30 mg/ 100mL (-) & (trace): 10 mL urine +2ml TCA : 0.5: < 10 mg/ 100 mL (1+): None : 1: 100-500mg/ 100ml (+2) & (+3): 2mL urine +4mL distl. H20 : 3:> 500 mg/ 100 mL (+4): 1 mL Urine +10 mL disl. H20 : 11Note: If precipitation of proteins is very excessive, dilute urine 1+ 10PROTEIN PRECIPITATION:Pipette into centrifuge tubes:Urine Sample 5.0 mLTCA (1.2 mmol/L) 1.0 mL Mix, allow to stand for 10 mins at 20-25 deg C, then centrifuge for 10 mins. Discard clear supernatant and place centrifuge tube on filter paper for 5 mins. Remove the residual supernatant by means of filter paper. Procedure: Wavelength: Hg 546 nmCuvette: 1 cm. light pathTemperature : 20-25 degCMeasure against solution 1.Pipette onto precipitate: Solution 1 : 5.0 mLDissolve while shaking gently, let stand for 30 mins. at 20-25 deg C, and read absorbance of sample (A sample) against solution 1 (reagent blank).Determination via Standard:If measurements cannot be carried out at Hg 546 nm, determine sample A in the range 530-570 nm and run additional standard. Mix 0.1 mL solution 1 and read as standardA after 30 mins. Calculation of the Concentration (c) Total Protein in the Sample::: Hg 546 nm (factor): 530-570 nm Standard: mg/ 100 Ml: C= 373 x F x Sample A: C= 118 x F x Sample A/Std AFor 24 hours Urine Computation:373 x Total Volume x Sample A x F (Dil. Factor) Gms/ 24 hrs = 1000/ 100Limitation of Procedure:A= 0.500 (Hg 546 nm)Source: Boehringer Mannheim GmbH Diagnostics

MicroalbuminMethod Used: Micral Test (Boehringer Manheim)Applications: For early detection and monitoring the course of incipient nephropathy (e.g. Diabetics and Hypertensive patients).Microalbumin denotes an albumin excretion of 20- 200 mg/LSample Material: It is recommended to use first morning urine, it is advisable to collect morning urine samples on three days of a given week and to analyze them within that week. Procedure:1. Dip the strip into the urine for 5 seconds up to just beneath the blue test zone. When dipping and withdrawing the strip, do not allow the strip to touch the side of the urine vessel.2. Place the strip on a non-absorbent surface across the top of the urine vessel.3. After 5 mins, compare the reaction colour with the colour scale on the test strip vial label, making sure that predominant colour on the test zone is assessed and not any other smaller patch of varying colour. Consequences of a Positive Test Result:The result is positive when at least 2 of 3 morning urines tested procedure a reaction colour corresponding to 20 mg/L albumin or more.

Urine Pregnancy TestMethod Use: Abott Test PackName and Intend Use:ABBOTT Test Pack plus hCG-Urine is a self performing immunoassay designed for the qualitative determination of human chorionic gonadotropin (hCG) in urine for early detection of pregnancy. Summary and Explanation of Test:Human chorionic gonandotropin (hCG) is a hormone secreted by the developing placenta shortly after fertilization. The appearance and rapid rise in the concentration of hCG in the mothers urine make it an excellent marker for confirming pregnancy. The ABBOTT Test Pack Plus hCG-Urine is a self performing immunoassay which utilizes both monoclonal and polyclonal antibodies to selectively identify hCG in urine with a high degree of sensitivity. Elevated levels of hCG can be detected before a first missed menses without interference from other hormones. Reagents:1. Remove the ABBOTT Test Pack Plus hCG-Urine Reaction disc from the pouch and place on a flat, dry surface. Use the transfer pipette supplied to dispense 3 gtts of specimen (180 uL) into the sample well.2. The test result should be read immediately after the appearance of a red color in the end of Assay Window (in approx.. 5 mins). This is important because specimen containg low levels of hCG may show color development over time. Interpretation of Results1. A positive (+) indicates that the specimen contains elevated levels of hCG. Any color on the patient bar should be interpreted as a positive result even if it has less color than the control bar. 2. A negative (-) indicates the absence of detectable hCG3. If a positive (+) or negative (-) sign fails to appear in the result window, or if now color is visible in the end of Assay Window, the specimen should be rested.4. With weak positive results it is good laboratory practice to resample and retest after an additional 48 hours.

Routine FecalysisI. Gross ExaminationA. Color: Brown: Dark Brown: Pale Brown: Yellow: Green: Clay: Others (specify)B. Consistency : Hard (resist puncture): Formed (can be punctured): Soft (can be cut): Loose (Assumes shape of the container): Diarrheic (flows): Water (pours)C. Blood: Absence or PresenceD. Mucus: Absence or PresenceII. Chemical Examination:A. Blood (occult) : Use Hematest. Report as follows:a. Negativeb. Weakly Positivec. Strongly PositiveIII. Microscopic ExaminationPrepare a direct saline fecal smear on one end of the slide and a direct iodine- stained fecal smear on the other end of the same slide.A. Direct Saline Fecal Smear:1. Place one drop physiologic saline on a slide.2. With an applicator stick, select a 1 mg fecal sample. Avoid selecting non fecal elements unless schistosome eggs or amoeba are suspected, in which case select flocks of mucus and blood. 3. Stir into saline and make a homogenous suspension. Remove coarse fibers, seeds, sands etc4. Cover with 22x22 mm. coverslip. If of proper consistency, the print of a newspaper will be just legible through the smear after applying the coverslip.

5. A satisfactory smear contains a maximum of observable fecal elements without any objects or protozoan size (8-30 uL in diameter) being obscured. IF the preparation is too dilute or too concentrated, discard it and prepare another.B. Direct Iodine- Stained fecal Smear: Dobell and OConnors1. Place one drop of iodine on a slide. (Iodine solution)2. Proceed as describe under item (A).Examination of Smear:The fecal smear should be examined under LPO magnification. A magnification of 100x is ample for detecting the presence of protozoan cysts or helminth eggs. After detecting suspicious objects, examine closely for more morphologic detail. Do not look for fecal parasites routinely with 430x. The fecal smear should be examined systematically. Start at the lower left edge of the smear and examine preparation up 1 field and move back toward the left. Examine the entire preparation in this left to right, right to left manner. Preparation of Lugols SolutionPotassium Iodide2.0 gramsPowdered Iodine Crystals1.0 gramsDistilled Water 100mL Stable up to 2 weeks if stored in a stoppered brown bottle.

Concentration ProcedureMethod: Formalin-EtherReference: Clinical Diagnosis and Management by Laboratory Method (Todd, Sanford, and Davidsohn), Vol II pp. 1737-1738Note: This procedure is efficient in recovering most protozoan cysts and helminth eggs and larvae, including operculate eggs and is moderately effective for schistosome eggs. Less distortion of cysts occurs with this technique than with this technique than with Zinc Sulfate method. Procedure: 1. Comminute a portion of a stool about 2 cm diameter in sufficient saline so that 10mL of suspension will yield about 1 mL of sediment upon centrifugation. 2. Strain about 10 mL of suspension through a small funnel containing wet gauze into a 15mL conical centrifuge tube.3. Centrifuge at 3200 rpm for 1-2 mins. Decant supernatant. About 1-1.5 mL of sediment should be present. If the amount is much larger or smaller, adjust to the proper quantity in the following manner:a. Amount too largeResuspend the sediment in saline and pour out a portion. For example, if the amount is twice the desired quantity, pour out half of the suspension and then add saline to bring the fluid level to about 10mL and centrifuge again.b. Amount too smallPour off the supernatant and strain a second portion of the original fecal suspension into the tube. The amount to be strained can be determined from the amount of sediment, that is, if about of the quantity necessary to obtain in the first centrifugation, strain another 10mL into the tube. Centrifuge again. It is not necessary to have an exact quantity of sediment in the tube, but the quantity should approximate the amount indicated. 4. Resuspend the sediment in fresh saline, centrifuge and decant as before.5. Add about 9 mL of 10% Formalin to the sediment, mix thoroughly, and allow to stand for 5 mins or longer. 6. Add 3 mL of Ether, stopper the tube and shake vigorously in an inverted postion for at least 30 seconds.7. Centrifuge at 2400 rpm for 1 min. Four layers should result as follows: (1) Layer of Ether; (2) Plug of Debris; (3) Layer of Formalin; (4) Sediment. 8. Free the plug of debris from the sides of the tube by ringing with an applicator stick and carefully decant and discard the top three layers. Use a cotton swab to clean debris from the walls of the tube.9. With a pipet, mix the sediment with a small amount of fluid that drains back from the slides of the tube and prepare iodine and unstained mounts for microscopic examination. If not enough fluid is left in the tube, a drop of saline or 10% Formalin can be added to the sediment. Examination for Amoeba:Note: The consistency of the fecal specimen determines the manner in which it should be processed for examination for protozoa.A. Loose, diarrheic or Watery: Fecal specimen with this type of consistency primarily contain the trophozoite stage and therefore must be examined immediately, or typical trophozoite recognition. To facilitate examination, do a direct saline fecal smear.B. Formed or Soft: Fecal specimen with this type of consistency contains primarily the cyst stages. Process the specimen as follows:a. Do a Direct Saline Fecal Smear and a Direct Iodine-Stained Fecal smear. b. If the results are negative for amoebic cysts, do the Formalin-Ether Concentration Technique.

Examination for Pinworms(Scotch Tape Swab Technique)Reference: Gradwohls Clinical Laboratory Methods and Diagnosis, 7th Ed. Vol II, pp. 1759Note: The fecal material from around the anus is usally examined for diagnosis of Enterobius vermicularis (pinworm infection). Often Taenia eggs undetected by direct smear examination, are also found in perianal swabs. The fecal swab should be collected either late at night or early in the morning before bathing or defacation. Three or four samples, collected on alternate nights, should be examined before a negative diagnosis for pinworm is made.Procedure:1. Cut a 6 in. strip of Scotch tape and bend over the sides of a tongue depressor, sticky side out. (Frosted or opaque tape is not satisfactory. Tongue depressors that have been squared are easier to handle.)2. Holding the tape close to the sides of the applicator with a finger and a thumb, spread the buttocks to expose the anus and press the sticky tape surface tape surface against the right and left peri-anal folds.3. Place the tape, sticky side down, on a clean slide containing a drop of toluene or xylene. Be sure the section of the tape containing fecal debris comes in contact with the toluene. 4. Search for pinworm eggs under the low power magnification.

Semen AnalysisPrinciple: Semen analysis involves gross examination (volume, color, turbidity, viscosity and pH) and microscopic examination (motility and spermatozoa count).Reagent: Spermatozoa fixative solution: Add 5 gms. Sodium Bicarbonate (NaHCO3) and 1 mL formalin to a 100 mL volumetric flask. Dilute to mark with distilled or cold water. Collection Instruction: A physician will usually give the instruction. However, the patient should be reminded of several critical points.1. The patient may be required to abstain from intercourse for a period directed by the physician. 2. The specimen should be collected in a clean container that has been prewarmed to body temperature. 3. The specimen should be delivered to the lab within 30 mins4. The specimen should be kept at body temperature not be subjected to extreme cold or heat.Gross Examination:1. Record the time of collection and receipt of specimen.2. Measure and Record the volume3. Measure and Record the following: Color (pearly white, yellow, etc)Viscosity (Viscous, Non viscous, Gelatin, Liquid)4. Determine the pH with a pH reagent strip and record this.Motility Examination:1. When the specimen becomes fluid (15 to 30 mins after collection the semen liquefies by the action of fibrinolysin, (place one drop on a slide (prewarmed 37 deg C) and place a coverslip on it. Record and Report the time of liquefaction.2. Under HPO, count the number of motile and non motile spermatozoa in 2 or more areas. Only those which moves forward actively are considered motile. Record the % of motile spermatozoa seen. Record any WBC or RBC seen.Spermatozoa Count:1. Draw the liquefied semen to the 0.5 mark of WBC pipet.2. Draw fixative solution to the 11 mark on the WBC pipet.3. Let the mixture stand until the mucus dissolves.4. Shake the pipet thoroughly and charge a hemacytometer5. Count the spermatozoa in the same manner as you would count WBC6. Calculations: No. of Sperm Counted x Dilution (20)_____ Sperm/ cu.mm= Sperm/ cu. mm x 1000Sperm/ mL= sperm/ cu.mm x 1000Differential Count:1. Make a smear, heat dry, and stain in hematology.2. Examine the morphology and report the % of abnormal forms. (Do a differential count up to 100 spermatozoa, identifying the normal and abnormal forms). Morphologically normal sperm are quite uniform in appearance. Any sperm with rounded, enlarged, small or bilobed heads are abnormal. Abnormal tails are enlarged, small, irregular in length and or multiple. Normal Values: Volume: 1.5 5.0 mLpH: greater than 7Motility: greater than 60%Sperm Count: 60 to 150 million/mL