CLINICAL CHEMISTRY III LABORATORY REPORT

MIDTERMS

ALDOSTERONE

Alegria, Neil Jenkin M., Casañas, Dennielle G., De Jesus, Earl James Leonard P., Lopez, Katrina May E., Lee, Kristine

Joy G., & Rabe, Yasmin Clarice C.

Department of Medical Technology, Institute of Arts and Sciences, Far Eastern University, Morayta, Manila 1108,
Philippines

ARTICLE INFO

Article history:
Date Performed: August 7, 2018 Aldosterone, the key hormone in the mineralocorticoid pathway, plays a
Date Submitted: August 27, 2018 fundamental role in salt and water homeostasis, blood pressure regulation,
and cardiovascular remodeling (Xanthakis & Vasan, 2014).

1.0 INTRODUCTION

Aldosterone is secreted by the outer portion of the adrenal glands, particularly from the zona glomerulosa. This

hormone is regarded as the principal regulator of the salt-and-water balance of the body. It aids in retaining sodium and

water within the body by binding to mineralocorticoid receptors and the mineralocorticoid-aldosterone complexes in the

cytoplasm of renal tubular cells which then increases the excretion of potassium in the body. Aldosterone has also minimal

effects in fat, carbohydrate, and protein metabolism (Stiefel et al., 2011).

Aldosterone is regulated by the renin-angiotensin-aldosterone-system. Renin which primarily released by the

juxtaglomerular cell of the kidneys which is stimulated by sympathetic nerve activation, renal artery hypotension, and low

sodium levels. Renin then acts on the substrate angiotensinogen forming angiotensin I. The angiotensin converting enzyme

from the lungs will act on angiontensin I to form angiotensin II stimulating the release of aldosterone (Goodfriend, 2007).

2.0 OBJECTIVES

• To measure the amount of aldosterone in the blood.

• To measure the amount of aldosterone removed in urine in a day.

• To correlate the results of the tests done with the fluctuation, or lack thereof, of aldosterone in the body

• To determine if there is a presence of any discrepancies in the patient sample

• To make a comparison between the patient’s values and the reference range

Professor Annlyn Sanchez Page 1

3.0 METHODOLOGY

3.1 Materials and Reagents

• Venipuncture Kit

• EasyLyte Plus

• Centrifuge

3.2.1 Procedure

A. ALDOSTERONE

Collect blood sample from the patient (Fig A.1) and transfer to red top non additive tube. Centrifuge the blood

samples for 5 minutes at 3000 RPM (Fig A.2). Gently remove, the blood sample from the centrifuge, to avoid mixing of the

serum and plasma.

Follow the instructions displayed in the screen. Align the tube vertically below the EasyLyte Plus needle-like

aspirator. It is a must to place the end of the aspirator at the middle of the serum portion of the blood. Do not remove the

tube until the aspirator goes back to the machine (Fig A.4). Wait until the results popped out of the screen. The screen will

display the serum’s sodium, potassium, and chloride (Fig A.5).

Figure A.1 Blood extraction Figure A.2 Centrifugation Figure A.3 Blood sample after
centrifugation.

Figure A.4 EasyLyte Plus Figure A.5 EasyLyte Plus reading

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4.0 RESULTS AND DISCUSSION
A. ALDOSTERONE

Aldosterone is a type of steroid hormone that acts primarily in renal collecting ducts to stimulate reabsorption of

Na+ as well as secretion of K+ and H+ (Muto, 1995). According to Endocrine society (2018), It also affects the body's ability

to regulate blood pressure. It sends the signal to organs, like the kidney and colon, that can increase the amount of sodium

the body sends into the bloodstream or the amount of potassium released in the urine. The hormone also causes the

bloodstream to re-absorb water with the sodium to increase blood volume. All of these actions are integral to increasing and

lowering blood vessels. Indirectly, the hormone also helps maintain the blood's pH and electrolyte levels.

In this experiment, aldosterone level was indirectly determined by means of the values of the electrolytes of the

patient’s serum sample. Sodium, potassium and chloride values of the sample was determined by means of EasyLyte Plus.

The following are the results:

Electrolyte mEq/L (Sample) mEq/L (Normal Value)

Sodium 142.1 136 – 145

Potassium 3.06 3.5 – 5.1

Chloride 104.4 98 – 107

Based on the reference values of the electrolytes from Bishop, Fody & Schoeff (2013), the electrolytes: sodium,

potassium and chloride of the patient’s blood sample was all in the normal range. This means that reabsorption of sodium

of the patient and its secretion of potassium and hydrogen ion are all metabolically normal. In addition, this results may

correlate in the patient’s blood pH and the body’s blood pressure.

Figure A.6 Renin Angiotensin Aldosterone System

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REFERENCES

Bishop, M. L., Fody, E. P., & Schoeff, L. E. (2013). Clinical Chemistry Principles, Techniques, and Correlations.

Philadelphia: Lippincott Williams & Wilkins.

Endocrine Society. (2018). What is aldosterone. Retrieved from https://www.hormone.org/hormones-and

health/hormones/aldosterone

Goodfriend, T. (2007). Aldosterone—A Hormone of Cardiovascular Adaptation and Maladaptation. Retrieved from

https://onlinelibrary.wiley.com/doi/abs/10.1111/j.1524-6175.2006.05110.x

Muto, S. (1995). Aldosterone. Retrieved from https://www.ncbi.nlm.nih.gov/pubmed/7743155

Stiefel, P., Vallejo-Vaz, A. J., García Morillo, S., & Villar, J. (2011). Role of the Renin-Angiotensin System and Aldosterone

on Cardiometabolic Syndrome. International Journal of Hypertension, 2011, 685238.

http://doi.org/10.4061/2011/685238

Strasinger, S. K., & Di Lorenzo, m. S. (2014). Urinalysis and Body Fluids. Philadelphia: F.A. DAVIS COMPANY.

Xanthakis, V., & Vasan, R. S. (2013). Aldosterone and the Risk of Hypertension. Current Hypertension Reports, 15(2),

102–107. http://doi.org/10.1007/s11906-013-0330-y

Professor Annlyn Sanchez Page 4

 CLINICAL CHEMISTRY III LABORATORY REPORT

MIDTERMS

ORAL GLUCOSE TOLERANCE TEST

Alegria, Neil Jenkin M., Casañas, Dennielle G., De Jesus, Earl James Leonard P., Lopez, Katrina May E., Lee, Kristine

Joy G., & Rabe, Yasmin Clarice C.

Department of Medical Technology, Institute of Arts and Sciences, Far Eastern University, Morayta, Manila 1108,
Philippines

ARTICLE INFO

Article history:
Date Performed: August 7, 2018
Date Submitted: August 27, 2018
Philippines is one of the 22 countries and territories of the IDF WP
(International Diabetes Federation Western Pacific) region. 425 million
people have diabetes in the world and 159 million people in the WP region.
There were over 3.721.900 cases of diabetes in the Philippines last 2017
with a percentage of 6.2 prevalence of diabetes in adults.

1.0 INTRODUCTION

Glucose is the most predominant monosaccharide. It is synthesized in the liver and kidneys from non-carbohydrate

intermediates (i.e. pyruvate or glycerol). It is an essential nutrient needed by the body for the production of energy. This

sugar is often found in ripen grapes, honey, and most of the sweet fruits hence its name “grape sugar” or dextrose which is

derived from its configuration, dextrorotatory. (Kullabs, 2017). Glucose is referred to as an aldohexose. It contains six carbon

atoms with a terminal aldehyde group on its terminal chain. It can either be in acyclic (chain) form or in cyclic (ring) form

(National Center for Biotechnology Information, 2018).

2.0 OBJECTIVES

• To evaluate apparent insulin release and insulin resistance in various clinical settings,

• To evaluate glucose metabolism,

• To correlate the results of the tests done with the fluctuation, or lack thereof, of glucose in the body,

• To determine if there is a presence of any discrepancies in the patient sample;

• To make a comparison between the patient’s values and the reference range.

Professor Annlyn Sanchez Page 1

3.0 METHODOLOGY

3.1 Materials and Reagents

• Venipuncture Kit

• Test tubes

• Dextrose powder

• Pipette/ Micropipette

• Centrifuge

• Water bath

• Spectrophotometer

3.2.1 Procedure

A. ORAL GLUCOSE TOLERANCE TEST

Dissolve 75 grams dextrose powder in a warm cup of water (Fig A.1). Collect blood sample from the patient (Fig

A.2) and transfer to red top non additive tube. Repeat blood extraction after one hour and two hours. The first blood collected

was kept into a refrigerator. Centrifuge the blood samples for 5 minutes at 3000 RPM. Gently remove, the blood sample

from the centrifuge, to avoid mixing of the serum and plasma.

Place the glucose working reagent in a 37oC water bath for five minutes. With the use of a micropipette, add one

mL of the reagent and ten uL of the sample in each of the three tubes (Fig A.4). Place in a water bath for three minutes (Fig

A.5). Read the absorbance of the standard, unknown and sample at 500nm. (Fig A.6).

Figure A.1 Dextrose powder Figure A.2 Blood extraction Figure A.3 Centrifugation

Figure A.4 Test reagent with serum Figure A.5 Water bath Figure A.6 Reading of the
spectrophotometer

Professor Annlyn Sanchez Page 2

4.0 RESULTS AND DISCUSSION
A. ORAL GLUCOSE TOLERANCE TEST

Oral glucose tolerance testing (OGTT) evaluates the ability to regulate the glucose metabolism. Plasma glucose

reflects a combination of carbohydrate uptake from the gastrointestinal tract, hepatic glucose uptake, and peripheral glucose

uptake. Oral GTT is currently the standard test for the diagnosis of diabetes. Random plasma glucose of greater than

200mg/dl, fasting blood glucose of greater than 126mg/dl, and 2 hours post prandial of greater than 200mg/dl using an oral

load of 75grams of anhydrous glucose dissolve in water are indicative of diabetes mellitus (Bartoli, 2008).

Oral GTT is unnecessary if fasting or random blood glucose levels are within the normal range. Indications for oral

GTT include unclear fasting plasma or random plasma glucose results, to screen for gestational diabetes mellitus at 24 to

28 weeks of gestation in all pregnant women not known to have diabetes, to screen for diabetes mellitus at 6 to 12 weeks

postpartum in women with a history of gestational diabetes mellitus, using non-pregnant oral GTT criteria and to screen for

impaired glucose tolerance, which is associated with an increased risk of developing diabetes mellitus.

Interpretation of the results is based on the fasting and 2-hour plasma glucose values after the administration of

75g glucose load within 5 minutes. There are requirements in performing the OGTT and this includes consulting doctors

regarding the medications the patient is currently taking for some medications can interfere with the results; abstaining from

food intake for at least 8 hours; and following the instructions strictly. Errors in patient preparation, type of sample

used/incorrect sample, and glucose load not consumed within the specified time (5 minutes) can give erroneous results.

A spectrophotometer is capable of displaying both transmittance and absorbance. Usually you will be required to

record one of these. Absorbance is the amount of light absorbed by a sample. The amount of monochromatic light absorbed

by a sample is determined by comparing the intensities of the incident light and transmitted light. The ratio of the intensity

of the transmitted light to the intensity if the incident light is called transmittance.

In this experiment the spectrophotometer was not able to give the absorbance of the sample, instead the

transmittance of the sample was generated. The absorbance of the sample was derived from Beer’s law

Figure A.6 Beer’s Law

Based from the given formula the resulting absorbance of 1.112 was derived from the transmittance 7.7268. With

these, the value for glucose was determined, 95.04 mg/dL. The patient is said to be normal since it lands between the

normal range which is 70-105 md/dL.

Professor Annlyn Sanchez Page 3

REFERENCES

Butler, S., Khanlian, S., & Cole, L. (2001). Detection of Early Pregnancy Forms of Human Chorionic Gonadotropin by

Home Pregnancy Test Devices. Retrieved from http://clinchem.aaccjnls.org/content/47/12/2131

Chard, T. REVIEW: Pregnancy tests: a review, Human Reproduction, Volume 7, Issue 5, 1 May 1992, Pages 701–710,

https://doi.org/10.1093/oxfordjournals.humrep.a137722

Gnoth, C., & Johnson, S. (2014). Strips of Hope: Accuracy of Home Pregnancy Tests and New Developments.

Geburtshilfe Und Frauenheilkunde, 74(7), 661–669. http://doi.org/10.1055/s-0034-1368589

International Genetically Engineered Machine. (2018). Controls. Retrieved from

http://igem.org/wiki/images/f/fc/Controls.png

Kullabs. (2017). Glucose chemistry. Retrieved from https://www.kullabs.com/classes/subjects/units/lessons/notes/note

detail/8529

Moraes, Gisele Silva de, Cristovam, Rafael do Amaral, & Savaris, Ricardo Francalacci. (2011). Comparative analysis of

the accuracy of urinary hCG tests in vitro. Revista da Associação Médica Brasileira, 57(5), 516-522.

https://dx.doi.org/10.1590/S0104-42302011000500008

National Center for Biotechnology Information. PubChem Compound Database; CID=5793,

https://pubchem.ncbi.nlm.nih.gov/compound/5793 (accessed Aug. 27, 2018).

Sally Suba. Clinical Endocrinology and Toxicology with Drug Testing Laboratory Manual 2014

Professor Annlyn Sanchez Page 4

 CLINICAL CHEMISTRY III LABORATORY REPORT

MIDTERMS

PREGNANCY TEST

Alegria, Neil Jenkin M., Casañas, Dennielle G., De Jesus, Earl James Leonard P., Lopez, Katrina May E., Lee, Kristine

Joy G., & Rabe, Yasmin Clarice C.

Department of Medical Technology, Institute of Arts and Sciences, Far Eastern University, Morayta, Manila 1108,
Philippines

ARTICLE INFO

Article history:
Date Performed: August 14, 2018
Date Submitted: August 27, 2018
Pregnancy tests are widely used both by the public and by healthcare
professionals. All tests depend on the measurement of human chorionic
gonadotrophin (HCG) in urine. Other pregnancy-specific materials have
been proposed as pregnancy tests but none can better the sensitivity and
convenience offered by immunoassay of HCG (Chard, 1992).

1.0 INTRODUCTION

Pregnancy test is done to confirm whether or not one is pregnant. According to Gnoth & Johnson (2014), this detects

the presence of the dimeric hormone hCG or human chorionic gonadotrophin. It is secreted by trophoblasts which enhances

implantation of the blastocyst and the placenta maintaining the corpus luteum in the first trimester of pregnancy.

There are two ways to determine pregnancy. One utilizes urine as a sample the other uses a blood sample. The

test can either be qualitative or quantitative. A qualitative hCG test which answers whether the person is pregnant or not.
The quantitative hCG test on the other hand measures the exact amount of hCG present. Immunochromatography is the

principle involved with this test. The test kit contains a conjugate pad which contains a mouse monoclonal anti-hCG antibody

that is conjugated with a colloidal gold and a nitrocellulose membrane with a test and control line. When the specimen is

applied, hCG from the specimen binds on the anti-hCG-gold conjugate forming a complex. This test can never be wrong

with a 99 percent accuracy (Moraes et al., 2011).

2.0 OBJECTIVES

• to determine whether the level of human chorionic gonadotrophin (hCG) in urine is consistent with a woman being

pregnant.

• to diagnose pregnancy, and evaluate and manage problems during pregnancy.

• To determine if there is any amount of hCG in the blood that could signal the presence of malignancy.

Professor Annlyn Sanchez Page 1

 • To determine if there is a presence of any discrepancies in the patient sample

• To make a comparison between the patient’s values and the reference range

3.0 METHODOLOGY

3.1 Materials and Reagents

• Urine in a screw-capped container

• Pasteur pipette

• Pregnancy test kit cassette

• Tissue paper

3.2.1 Procedure

A. PREGANCY KIT CASETTE

Collect a mid-stream clean catch urine in a screw-capped container (Fig A.1). Cool the urine at room temperature.

Tear the packaging of the kit gently, avoid dropping the cassette (Fig A.2). Place the cassette on a clean tissue paper.

Aspirate urine by a Pasteur pipette (Fig A.3). A few centimeters above from the test kit, align the pasteur pipette on the

region near the “S” sign was, then add three drops of the urine (Fig A.4). Wait for about three minutes and read the results

(Fig A.5).

Figure A.1 Mid-stream catch urine Figure A.2 Cassette from the Figure A.3 Centrifugation
packaging

Figure A.4 Adding of sample to the Figure A.5 Cassette after three
cassette minutes of the administration of
the urine

Professor Annlyn Sanchez Page 2

4.0 RESULTS AND DISCUSSION
A. PREGNANCY KIT

Pregnancy test is used for the detection of the hormone, human chorionic gonadotropin (hCG) that is present when

an individual is pregnant. hCG is made by cells formed in the placenta, which nourishes the egg after it has been fertilized

and becomes attached to the uterine wall. It can be detected 12 to 14 days after conception by a urine test. Typically, the

hCG levels will double every 72 hours. The level will reach it’s peak in the first 8 to 11 weeks of pregnancy and the will

decline and level off for the remainder of the pregnancy. A low level hCG can indicate possible miscarriage or blighted ovum

and ectopic pregnancy. A high hCG levels can indicate molar pregnancy or multiple pregnancy (Butler et al., 2001).

In this experiment, a simple methodology was performed. A pregnancy kit from Abon Biopharm was used. The

packaging has instructions to be followed at the back. As the students performed the procedure, a single violet line showed,

as indicated in the kit’s packaging, a double violet line indicates a positive result, and single violet line is the indicator of

negative. In this procedure, the kit showed a single prominent violet line which indicated that the patient was negative.

Figure A.6 Demonstrates the positive and negative control of a pregnancy kit

Professor Annlyn Sanchez Page 3

REFERENCES

Butler, S., Khanlian, S., & Cole, L. (2001). Detection of Early Pregnancy Forms of Human Chorionic Gonadotropin by

Home Pregnancy Test Devices. Retrieved from http://clinchem.aaccjnls.org/content/47/12/2131

Chard, T. REVIEW: Pregnancy tests: a review, Human Reproduction, Volume 7, Issue 5, 1 May 1992, Pages 701–710,

https://doi.org/10.1093/oxfordjournals.humrep.a137722

Gnoth, C., & Johnson, S. (2014). Strips of Hope: Accuracy of Home Pregnancy Tests and New Developments.

Geburtshilfe Und Frauenheilkunde, 74(7), 661–669. http://doi.org/10.1055/s-0034-1368589

International Genetically Engineered Machine. (2018). Controls. Retrieved from

http://igem.org/wiki/images/f/fc/Controls.png

Kullabs. (2017). Glucose chemistry. Retrieved from https://www.kullabs.com/classes/subjects/units/lessons/notes/note

detail/8529

Moraes, Gisele Silva de, Cristovam, Rafael do Amaral, & Savaris, Ricardo Francalacci. (2011). Comparative analysis of

the accuracy of urinary hCG tests in vitro. Revista da Associação Médica Brasileira, 57(5), 516-522.

https://dx.doi.org/10.1590/S0104-42302011000500008

National Center for Biotechnology Information. PubChem Compound Database; CID=5793,

https://pubchem.ncbi.nlm.nih.gov/compound/5793 (accessed Aug. 27, 2018).

Sally Suba. Clinical Endocrinology and Toxicology with Drug Testing Laboratory Manual 2014

Professor Annlyn Sanchez Page 4