There are several medical issues that are of unique concern to women. 

Now your gynecologist can order four women’s health care tests in one cytology collection for:

1. Sequencing HPV assay
2. Sequencing BRCA1 and BRCA2 Ashkenazi Jewish founder gene mutations screen
3. Sequencing Chlamydia trachomatis test
4. Sequencing Neisseria gonorrhoeae (GC) test

   The residual cells (about 10%) in a non-contaminated SurePath or ThinPrep vial after the Pap cytology screen can be used for all four tests. The target DNA is stable in the fixatives for at least 6 months at ambient temperature.  Our positive test reports always contain a DNA sequence electropherogram, similar to an electrocardiogram (EKG) in a cardiologist’s consultation.

To request a test, download and fill out the form here.

BRCA1 and BRCA2 Mutations Screening

   Mutations in the BRCA1 and BRCA2 genes have been linked to certain types of hereditary breast and ovarian cancers.  About 10% of cases of ovarian cancer and 3–5% of cases of breast cancer are due to mutations in these genes.  Occurrence of mutation is about 1 in 300 and 1 in 800 for BRCA1 and BRCA2 genes, respectively, and are found more frequently in certain ethnic groups, including people of Ashkenazi (Eastern European) Jewish, French Canadian, and Icelandic backgrounds, but they can occur in anyone.  Inheriting one of these mutations increases the risk of getting breast cancer, ovarian cancer, and other types of cancer. 

   For more information, please see the education fact sheet from the American Congress of Obstetricians and Gynecologists (ACOG).

   Milford Molecular Diagnostics Laboratory is the only laboratory that has been certified according to the Clinical Laboratory Improvement Amendments (CLIA) requirements to perform the BRCA1 and BRCA2 Ashkenazi Jewish founder gene mutations screen on liquid-based cytology sample (SurePath or ThinPrep).

Why Sanger sequencing electropherograms should be attached to a report of BRCA1 and 2 mutations screen?

In this era of precision medicine, it is important to have physical evidence in the laboratory report for the detection of a wild type, a homozygous mutant or a heterozygous mutant in the BRCA1 185delAG, BRCA1 5382insC and BRCA2 6174delT locations. We have developed the first testing procedure to perform Sanger sequencing-based BRCA 1 and 2 Ashkenazi founder mutation screen on routine Pap cytology samples. ¹⁴

The following 3 sequencing electropherograms show a negative BRCA1 185delAG mutation (wild-type), a homozygous BRCA1 185delAG mutation, and heterozygous BRCA1 185delAG mutation from top to bottom.

The following 3 sequencing electropherograms show a negative BRCA1 5382insC mutation (wild-type), a homozygous BRCA1 5382insC mutation, and heterozygous BRCA1 5382insC mutation from top to bottom.

The following 3 sequencing electropherograms show a negative BRCA2 6174delT mutation (wild-type), a homozygous BRCA2 6174delT mutation, and heterozygous BRCA2 6174delT mutation from top to bottom.

Why sequencing human papillomavirus (HPV) genotyping is important in precision women’s health care?

   Most HPV infections of the female cervix are transient, self-resolving and harmless. Only when there is evidence of persistent infection by certain (high risk) HPV genotypes, the risk of developing cervical cancer is increased. This is based on nested PCR and DNA sequencing research, using technology similar to what we are using here to perform the studies. ¹⁷ There are more than 200 HPV genotypes, subtypes and variants have been reported. But we have recorded more than 40 HPV genotypes in southern Connecticut. Some of the HPV genotypes or variants in Connecticut have not reported in the world literature and probably cannot be diagnosed by the commercial test kits. One of such cases is illustrated below. We call it HPV-39 Milford variant.

(Published by Lee SH, et al. HPV infection among women in a representative rural and suburban population of the USA. Int J Gynaecol Obstet. 2009;105:210-4.)

This patient had 2 HPV assays by a commercial laboratory reporting questionable “positive for low risk HPV DNA and high risk HPV DNA” before December 2007 when a DNA sequencing HPV test was performed, showing a signature sequence for HPV-39 with a base mutation from “A to C” (arrow; compared with sequence GenBank Locus U45905), as shown in the upper electropherogram [12-2007]. Five months later, a follow-up sequencing HPV assay showed an HPV-39 L1 DNA with the same “A to C” mutation [05-2008], indicating a new variant of HPV-39 in which the 14th amino acid of the L1 capsid protein has permanently changed from leucine (codon tta) to valine (codon gta, read as TAC in these tracings), causing possible persistent HPV infection. Patients with persistent (same-genotype HPV) HPV infections, rather than consecutive transient infections by different HPV types, should be closely monitored with cytology Pap smears.

Why is a highly sensitive nested PCR needed for cervical HPV screening?

The commercial HPV test kits were designed to screen for CIN1/CIN2 precancerous cells which usually contain hundreds to thousands of copies of episomal HPV DNA per cell. However, as carcinogenesis progresses, the truly malignant cells, such as the well-known SiHa cancer cells, may contain a single copy of HPV16 integrated in the chromosome.20 Nested PCR is usually needed17 for detection of the limited copies of HPV in the malignant cells of invasive cervical cancer or cervical carcinoma in-situ. The nested PCR method used in Milford Molecular Diagnostics can even detect a very minute amount of residual HPV L1 gene DNA bound to aluminum adjuvant in an HPV vaccine. 21, 22

The concern of false negatives in HPV screening is shared by the College of American Pathologists in a message sent to its members on September 19, 2017 as follows:

For more information regarding the cited research, please visit our REFERENCES page to find details behind the footnote references.