Molecular Genetics Platelet Disorders Panel

Inherited platelet disorders are a group of genetic conditions that cause excessive and uncontrollable bleeding. These life-threatening conditions cause complications following surgery, childbirth, trauma or internal hemorrhages. Pathogenic variants in genes encoding cytoskeletal proteins, platelet adhesions and signaling pathways triggering coagulation are analyzed in this next-generation sequencing-based panel test. The test also includes genes associated with Bernard-Soulier syndrome, Scott syndrome, Glanzmann’s thrombasthenia, Thromboxane A2 receptor deficiency, GP6 collagen receptor deficiency, and Hermansky-Pudlak syndrome. The mode of inheritance for platelet disorders is autosomal recessive or autosomal dominant. The overall incidence for inherited platelet disorders is estimated to be ~1:10,000. 

Reasons for Referral:

• Abnormal bleeding or bruising
• Uncontrollable bleeding
• Positive family history for bleeding disorders (targeted testing is available if familial mutation is known.)
• Carrier testing.

Next generation sequencing will analyze the exons or coding regions of 71 associated genes with platelet disorders using Illumina NextSeq 500/550 technology. Samples are prepared using hybridization probes to enrich exonic regions.  This assay does not assess regions of insufficient coverage, introns and promoter regions; pseudogenes; where the reference genome is inaccurate or contains gaps and insertions; and regions of high GC or polynucleotide repeats, but may contain variants that impact gene function.

Exon-level deletion/duplication analysis is performed by running the NGS data through the Genome Analysis Toolkit (GATK) Germline Copy Number Variation best practices pipeline from GATK, version 4.1.4.1. A Bayesian model was validated clinically in our lab. The model can detect copy changes at a resolution of three (3) or more probe targets (exons) for deletions and duplications in genes that do not have pseudogenes, and is not designed to detect low-level mosaicism or balanced alterations.

Coding region coverage for the Platelet Disorders panel genes

Gene Name	Coverage
A2M, ABCA1, ANKRD26, ANO6, AP3B1, BLOC1S6, CD36, DHCR24, DPAGT1, DTNBP1,F2, F7, F8, F9, F10, F11, F13A1, F13B, FERMT3, FGA, FGB, FGG, FLNA, GATA1, GFI1B, GGCX, GNAS, GP1BA, GP6, HPS1,HPS3, HPS4, HPS5, ITGA2, ITGA2B, ITGB3, KLKB1, LMAN1, LYST, MASTL, MCFD2, MLPH, MPL, MYH9, MYO5A, NBEAL2, P2RX1, P2RY12, PLA2G4A, PLA2G7, PLAU, RAB27A, RASGRP2, SERPINA1, SERPINE1, SERPINF2, STIM1, TBXAS1,USF1, VIPAS39, VKORC1, VPS33B, VWF*, WAS	95%-100%
HPS6, TBXA2R, GP9, GNAQ, HOXA11, RUNX1	75%-95%

Coding regions below 95% may be covered upon request if further assessment of a gene is necessary (some exceptions may apply). Please contact Knight Diagnostic Laboratories for more information.

*VWF Exon 26 (NM_000552.3), a region with high homology is assessed via long-range PCR & Sanger sequencing

Blood:  EDTA or ACD (Solution A or B):

• Adult: 5 mL
• Child: 5 mL
• Infant: 2-3 mL

Saliva: 2 ORAgene™ Saliva Collection Kits (OGR-500) used according to manufacturer instructions.  Please contact KDL Client Services for a Saliva Collection Kit for patients that cannot provide a blood sample.

Assisted Saliva: 4 ORAgene™ Assisted Saliva Collection Kits (OGR-575) used according to manufacturer instructions.  Please contact KDL Client Services for an Assisted Saliva Collection Kit for patients that cannot provide a blood sample.

Skin Fibroblast: Punch Biopsy (Cell cultures will be prepared at KDL and used for testing), or 2 T-25 confluent flasks.

DNA: 5-10µg at a minimum of 60-100ng/µL (DNA must be extracted in a CLIA-certified laboratory or a laboratory meeting equivalent requirements as determined by the CAP and/or CMS).

Prenatal:

• Direct Amniotic Fluid (10-20mL)
• Direct CVS
• Direct POC
• Cultured Amniocytes (2 T-25 flasks)
• Cultured CVS (2 T-25 flasks)
• Cultured Fetal Tissue: Product of Conception (2 T-25 flasks)
• Cord Blood (1-2mL)

Notice Regarding Molecular Genetic Testing Prenatal Specimens:

Maternal cell rule-out testing will be performed on all prenatal specimens received. Please provide maternal blood (or saliva) in addition to the fetal specimen. Additional charges apply for the maternal cell rule-out test.

For routine testing of blood and saliva (or DNA extracted from them), KDL does NOT accept samples from patients within two (2) weeks of a packed cell/platelet transfusion or within four (4) weeks of a whole blood transfusion.  For extraordinary circumstances, where testing must be performed outside of the above windows, please contact our lab.  

For patients with bone marrow transplants, the only acceptable specimen type is a skin biopsy (Cell cultures will be prepared at KDL before testing).

A REQUISITION FORM MUST ACCOMPANY ALL SAMPLES.  Please include detailed clinical information, including ethnicity, clinical history, and family history.

Weekly

6-8 weeks

• Package and ship specimen to remain cold, but not frozen. 
• Ship via overnight express, using the FedEx priority overnight label provided. 
• Contact Client Services for shipping kits and instructions at (855) 535-1522.

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2. Hematology Am Soc Hematol Educ Program. 2011;2011:377-383
3. Hematology Am Soc Hematol Educ Program. 2005:396-402