College of American Pathologists
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What the manufacturers offer for CF screening

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January 2003
William Check, PhD

Having a defined set of mutations to test has facilitated the development of commercial reagents for cystic fibrosis carrier screening.

“When the assay was homebrew, it was very difficult to set up,” says Wayne Grody, MD, PhD, of the University of California, Los Angeles, School of Medicine. “Once the recommendations came out, companies started making more robust products that most molecular diagnostic laboratories can perform and validate.”

CF screening assays are still homebrew in that reagents are not FDA-reviewed but are analyte specific reagents, or ASRs, and each laboratory has to validate its assay.

“Manufacturers were very interested during the process of selecting a mutation panel,” says Sue Richards, MD, of Baylor College of Medicine. “Now we are trying to keep them informed as we revise the panel, so they can rapidly revise their ASRs.”

Four vendors offer ASRs for CF screening, with two more in development. On the most recent CAP Survey, Dr. Grody says, the 50 laboratories distributed among the following methods (vendors making each type of assay are in parentheses):

  • Probe hybridization, 38 percent (Roche, Innogenetics)
  • Oligonucleotide ligation assay (OLA), 26 percent (ABI)
  • Restriction enzyme digestion and electrophoresis, 18 percent
  • Allele-specific PCR, 16 percent (Orchid)
  • DNA sequencing, 16 percent
  • Mutation scanning, eight percent
  • “Other,” 26 percent

Probe hybridization assays are popular because they don’t require a large piece of equipment. Roche’s ASR of this type is probably the most popular CF screening product. One person who chose Innogenetics’ version is Timothy Stenzel, MD, PhD, assistant professor of pathology and director of the molecular diagnostics laboratory at Duke University Medical Center. “My technologists preferred doing the strip-based method,” Dr. Stenzel says, “and they liked Innogenetics’ best. For laboratories already doing PCR-based testing, it would be easy to set up.”

David Witt, MD, of Kaiser Permanente Northern California, uses in-house multiplex PCR with probe hybridization. He pools five patient samples for each run. “With a carrier incidence of about one in 25, we are likely to get a normal result in most pools,” Dr. Witt says. For positive pools, each sample is re-run individually.

Elaine Lyon, PhD, of ARUP Laboratories, chose ABI’s OLA. “We have been very pleased with it,” she says, “although we are always looking at other technologies to see if we can reduce turnaround time and labor costs.”

A non-PCR method in development, Third Wave Technologies’ Invader platform, was selected by Ronald McGlennen, MD, of Access Genetics, as the basis of his company’s kit. “It is simple and should be applicable to a large number of laboratories,” he says. He also likes that probes are pooled in five batches. “On a population basis, 97 to 98 percent of samples will be negative [in CF screening],” Dr. McGlennen says. Positive samples can be sent to a reference laboratory.

Finally, Nanogen is developing an electronic microarray product that employs screening for the ACMG/ACOG-recommended mutations followed by genotyping of positive samples on the system. This technology analyzes the 5T/7T/9T region only when the relevant R117H mutation is present.

“There are many good choices out there,” says Dr. Richards.