Pregnancy Test And Pap Smear

“PREGNANCY TEST” • Misnomer - do not actually determine pregnancy, but human chorionic gonadotrophin (hCG) - used to diagnose conditions other than pregnancy

HUMAN CHORIONIC GONADOTROPHIN

Alpha polypeptide subunit MW= 18,000 Beta polypeptide subunit MW=

Bioassay • 1928 - Aschheim-Zondek: Also known as the A-

Z Test, this very first scientific pregnancy test was developed in Germany. It involved several subcutaneous injections of varying amounts of a woman's urine into the backs of immature female mice weighing between five and eight grams. One hundred hours after the first injection, the mice were killed and the ovaries were examined. The A-Z test is named for German researchers Selmar Aschheim (1878-1965) and Bernhard Zondek (1891-1966). Zondek was the first person to describe the ovarian hormone and earned worldwide renown for discovering gonadotropins.

• 1931 - Friedman: A refined version of the AschheimZondek test, the Friedman test used rabbits instead of mice. Its advantages were the availability of the animals, a reduced error rate, and the reduced time required for completion of the test. The subject's urine was injected into the ear vein of a female rabbit. The test, named for Maurice H. Friedman (1903-1991), could be read within 36 to 48 hours after injecting the urine.

•1939 - Hogben (Xenopus): In the Hogben test, a female African clawed toad (Xenopus laevis) is injected with urine (or an extract) into the dorsal lymph sac. The presence of five, six, or more eggs within four to twelve hours indicates pregnancy. A similar test was done using male frogs or male toads. A woman's urine or serum is injected into the dorsal lymph sac of two male frogs (Rana pipiens) or male toads (Bufo marinus). The presence of spermatozoa in the cloacal fluid of both animals is positive; in one animal, inconclusive; in neither animal, negative. This test is named for the British biologist Lancelot Hogben (1895-1975). Although the Hogben pregnancy test had the advantage of not killing the test animals, it was replaced by

Limitations: • • • • •

Interference from LH Technical difficulty Standardization High Cost Maintain animal colonies

Immunoassay • Started in 1960’s • Expressed in International Units • 1 IU = 83.3 mg/dl

Immunoassay Hemagglutination Inhibition (Urine) • Uses Anti-HCG • If HCG is present, Neutralization would occur • HCG coated RBC indicator cells would sink to the bottom of test tube and form a ring • Took 1 to 2 hours

Immunoassay Hemagglutination Inhibition (Urine)

Immunoassay Latex Particle Agglutination Inhibition

Immunoassay Direct Latex Particle Agglutination

• Tube tests HAI and LAI 1-2 IU (5-7 days after first missed menses • Slide tests 2-5 IU (14-21 days after missed)

Immunoassay Radioimmunoass ay

•Uses radioisotopes •More sensitive and could use serum •Not affected by LH as much •Need special equipment to read •Usually was done when early dx was important

Immunoassay Enzyme Immunoassay

ELISA Kit

Immunoassay Enzyme •Monoclonal Immunoassay Beta - HCG Enzyme Tests •Very sensitive •Can pick up .05 IU (1 week after implantation) • specificity is high because cross reaction with other hormones besides hCG is not a

Quantitative Pregnancy Testing • Ectopic pregnancies low levels • low levels in 1 st trimester = bad prognosis • Trophoblastic Tumors - Hydatiform Mole - benign grape-like mass - Choriocarcinoma - may follow usual childbirth or incomplete removal of hydatiform mole, malignant *Increased levels of up to 6,000,000 IU vs. 100,000 in normal pregnancy

• Increased HCG -Seen in multiple pregnancies and Eclampsia -High levels HCG seen in males that have testicular tumors

Home pregnancy test

Home pregnancy test • • • • • •

Wait at least 1 week after the date of the expected period Use first morning urine Try to test the urine sample immediately after collection If testing occurs at other times of day, restrict fluid intake for 4-6 hours Waiting 10 minutes to read results improve results If test is negative, test again in 1 week if menstruation has not started

Human Placental Lactogen (HPL) • • • •

Synthesized in large quantities by placenta during last two trimesters Structurally similar to both prolactin and growth hormone Insulin antagonist Exact role is unclear

Alpha Feto Protein (AFP) •Detection of Neural Tube Defects (NTDs) Neural tube defects include both spina bifida(improper closure of the fetal spine) and anencephaly(improper closure of the fetal skull). Anencephaly is a lethal condition.

Pap Smear Collection Technique

Who should have a Pap smear? • The American Cancer Society recommends that you have your first Pap smear about three years after first having sexual relations or at age 21. After age 21, the guidelines are as follows:

Prior to Pap Smear: Try to schedule your Pap smear when you are not menstruating. If your cycle is unpredictable, and you start your period when it will coincide with your appointment, call your doctor's office as soon as possible to see if the doctor may recommend rescheduling. To ensure that you get the most accurate results, it is recommended that 48 hours before having a Pap smear you should not: • • • • •

have sex douche use tampons use spermicidal foams, creams, or inserts use vaginal creams or suppositories All of these things can wash away or hide abnormal cells.

Positioning • • • •

PARENTAL GUIDANCE

Privacy Buttocks just off table Good Lighting Drape

Padded Stirrups • Soft, padded stirrups • Oven mitts • Socks

Inspect • • • •

PARENTAL GUIDANCE

Spread labia Discharge Ulcers Growths

Anatomy

PARENTAL GUIDANCE

Vaginal Speculum

Warm Speculum • • • •

Warm water Not too hot Lubricates speculum Don’t use K-Y, Surgilube or Vaseline to lubricate speculum

Insert Speculum • Spread labia • Keep labia apart • Blades remain closed until fully inserted

PARENTAL GUIDANCE

Squamo-Columnar Junction • Junction of pink cervical skin and red endocervical canal • Key portion of the cervix to sample • Most likely site of dysplasia

Ayers Spatula • Concave end to fit the cervix • Convex end for vaginal wall and vaginal pool scrapings

Sample Cervix • Use concave end • Rotate 360 degrees • Don’t use too much force (bleeding, pain) • Don’t use too little force (inadequate sample)

Cytobrush • Insert ~ 2 cm (until brush is fully inside canal) • Rotate only 180 degrees (otherwise will cause bleeding)

Make Pap Smear • As thin as possible • Properly labeled

Spray with Fixative • Within 10-15 seconds • Allow to fully dry before packaging • Cytologic Fixative (hairspray works acceptably also)

Factors That Diminish the Accuracy of Pap Smears • Clinician Related Factors - Contamination with blood or lubricants - Mislabeled or unlabeled slides - Inadequate clinical history - Inadequate sampling of the transformation zone - Slide material too thick or insufficient - Performing pap in spite of obvious infection • Laboratory Related Factors - Confusing smears or names - Failure to identify dysplastic cells - Misinterpretation of diagnostic cells - Poorly controlled technical process

Background • George Papanicolaou, MD, PhD, discovered that tumor cells could be found in vaginal fluid of women with cervical cancer • 1928 - presented his paper entitled "New Cancer Diagnosis" at the Third Race Betterment Conference in Battle Creek, Michigan, • 1943 - the concepts of early cancer and carcinoma in situ were widely understood, and the potential of the "Pap smear" for cancer prevention was finally appreciated

The Pap Smear

• It was a Canadian physician, J. Ernest Ayre, described the method we know today as the Pap smear • 1940s, cytology laboratories were opening, and by the 1950s, Pap smear screening was widespread, even before clinical trials could be performed

UNDERLYING PRINCIPLE OF CYTOLOGY EXFOLIATIVE CYTOLOGY

DIFFERENT TYPES OF CELLS ARE PERIODICALLY DESQUAMATED FROM

THE NORMAL CELLULAR ELEMENTS : SUPERFICIAL CELLS

INTERMEDIATE CELLS

NAVICULAR CELLS

PARABASAL CELLS

ENDOCERVICAL CELLS

ENDOCERVICAL CELLS

LACTOBACILLI / COCCOBACILLI

GARDNERELLA VAGINALIS (CLUE CELLS)

M

26

Candida spp.

Candida spp.

Leptothrix

ACTINOMYCES SPP. M

27

TRICHOMONAS VAGINALIS M

28

Herpes simplex

NORMAL

LSIL

LSIL

HSIL

CIN2

HSIL (Moderate dyskaryosis)

HSIL

CIN3

HSIL (Severe dyskaryosis)

SCCA

ASCUS

Atypical squamous cells of undetermined significance (ASCUS) • is defined, according to the Bethesda System, as squamous abnormalities that are more marked than those attributable to reactive changes, but that quantitatively or qualitatively fall short of a definitive diagnosis of a squamous intraepithelial lesion • cells as having nuclei about two to three times normal size with normochromatic or slightly hyperchromatic nuclei, even chromatin distribution, and smooth or only slightly irregular nuclear membranes.