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Peroxisomal Disorders Sequencing Panel

  • Summary and Pricing
  • Clinical Features and Genetics
  • Citations
  • Methods
  • Ordering/Specimens
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TEST METHODS

NGS Sequencing

Test Code Test Copy GenesCPT Code Copy CPT Codes
1979 ABCD1 81405 Add to Order
ABCD3 81479
ACOX1 81479
AGPS 81479
DNM1L 81479
GNPAT 81479
HSD17B4 81479
PEX1 81479
PEX10 81479
PEX11B 81479
PEX12 81479
PEX13 81479
PEX14 81479
PEX16 81479
PEX19 81479
PEX2 81479
PEX26 81479
PEX3 81479
PEX5 81479
PEX6 81479
PEX7 81479
PHYH 81479
Full Panel Price* $1690.00
Test Code Test Copy Genes Total Price CPT Codes Copy CPT Codes
1979 Genes x (22) $1690.00 81405, 81479(x21) Add to Order
Pricing Comment

If you would like to order a subset of these genes contact us to discuss pricing.

Targeted Testing

For ordering targeted known variants, please proceed to our Targeted Variants landing page.

Turnaround Time

The great majority of tests are completed within 28 days.

Clinical Sensitivity

Pathogenic variants in the 14 PEX genes account for nearly all cases of clinically and biochemically diagnosed PBD-ZSS. The overall sensitivity of this sequencing panel test is predicted to be over 95% for DNA substitutions (i.e. missense, nonsense, splicing mutations) and small insertions and/or deletions (i.e. frameshift and splice-site mutations). Large deletions are generally not detected by sequencing (Waterham and Ebberink 2012). PHYH pathogenic variants account for more than 90% of patients with Adult Refsum disease. PEX7-Related Adult Refsum disease is very rare, and less than 10 patients have been reported to date (Jansen et al. 2004). Over 93% of patients with X-ALD are caused by ABCD1 pathogenic variants which can be detected by sequencing. Gross deletions/duplications account for the remaining ~7% of causative mutations (http://www.x-ald.nl; Steinberg et al. 2012). PEX7 pathogenic variants account for over 90% of all individuals with Rhizomelic chondrodysplasia punctate. The remaining 10% are caused by defects in GNPAT and AGPS (Braverman et al. 2012). To date, only about 20 patients with RCDP type 2 and less than 10 patients with RCDP type3 have been reported (Thai et al. 2001; Itzkovitz et al. 2011).

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 ABCD1$690.00 81479 Add to Order
ACOX1$690.00 81479
AGPS$690.00 81479
GNPAT$690.00 81479
HSD17B4$690.00 81479
PEX1$690.00 81479
PEX10$690.00 81479
PEX12$690.00 81479
PEX13$690.00 81479
PEX14$690.00 81479
PEX16$690.00 81479
PEX19$690.00 81479
PEX2$690.00 81479
PEX26$690.00 81479
PEX3$690.00 81479
PEX5$690.00 81479
PEX6$690.00 81479
PEX7$690.00 81479
PHYH$690.00 81479
Full Panel Price* $1290.00
Test Code Test Copy Genes Total Price CPT Codes Copy CPT Codes
600 Genes x (19) $1290.00 81479(x19) Add to Order
Pricing Comment

# of Genes Ordered

Total Price

1

$690

2

$730

3

$770

4-10

$840

11-30

$1,290

31-100

$1,670

Over 100

Call for quote

Turnaround Time

The great majority of tests are completed within 28 days.

Clinical Sensitivity

Over 93% of patients with X-ALD are caused by ABCD1 pathogenic variants which can be detected by sequencing. Gross deletions/duplications account for the remaining ~7% of pathogenic variants (http://www.x-ald.nl; Steinberg et al. 2012). For other peroxisomal disorders, clinical sensitivity for del/dup testing is expected to be low. Gross deletions or duplications not detectable by sequencing have been reported in PEX1, PEX6, PEX13, PEX14, PEX16, PEX26, PHYH, AGPS, ACOX1, ABCD3, and HSD17B4 (Human Gene Mutation Database).

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Clinical Features

Peroxisomal disorders are genetically heterogeneous metabolic diseases, and are divided into peroxisomal biogenesis disorders and peroxisomal single protein defects. Peroxisomal biogenesis disorders (PBDs) are caused by defects in peroxisome assembly and formation. Clinical features and biochemical findings are associated with multiple abnormal enzymatic pathways in peroxisomes. PBDs include Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum (PBD-ZSS) and PEX7-related disorders (rhizomelic chondrodysplasia punctata Type 1 and adult Refsum disease). PBD-ZSS consists of three related diseases (Zellweger syndrome (ZS), neonatal adrenoleukodystrophy (NALD), and Infantile Refsum’s disease (IRD)) that share overlapping phenotypes with severity ranging from ZS to IRD (Steinberg et al. 2012). Peroxisomal single protein defects include X-linked adrenoleukodystrophy (X-ALD), PHYH-related adult Refsum disease, rhizomelic chondrodysplasia punctata type 2 and type 3, congenital bile acid synthesis defect-5, D-bifunctional enzyme deficiency, and acyl-CoA oxidase deficiency (also called pseudoneonatal adrenoleukodystrophy). X-ALD is the most common inherited peroxisomal disorder and affects 1 in 18,000 individuals (Wanders and Waterham 2006).

Genetics

This NextGen test analyzes multiple genes involved in both peroxisomal biogenesis disorders and peroxisomal single protein defects. Except in the case of the ABCD1 and DNM1L genes, peroxisomal disorders are autosomal recessively inherited and caused by loss-of function mutations in the relevant gene. X-linked adrenoleukodystrophy (ABCD1 gene) is inherited in an X-linked recessive manner. DNM1L-assocated diseases can be an autosomal dominant condition secondary to dominant-negative missense mutations (Waterham et al. 2007) or be a recessive disorder due to loss-of-function mutations (Yoon et al. 2014). Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum (PBD-ZSS): PEX1, PEX2, PEX3, PEX5, PEX6, PEX10, PEX11B, PEX12, PEX13, PEX14, PEX16, PEX19, PEX26, DNM1L X-linked adrenoleukodystrophy (X-ALD): ABCD1 Rhizomelic chondrodysplasia punctate (RCDP): PEX7, GNPAT, AGPS Adult Refsum disease (ARD): PHYH, PEX7 Congenital bile acid synthesis defect-5: ABCD3 D-bifunctional enzyme deficiency: HSD17B4 Acyl-CoA oxidase deficiency: ACOX1 See individual gene test descriptions for information on clinical features and molecular biology of gene products.

Testing Strategy

For this Next Generation (NextGen) panel, the full coding regions plus ~20 bp of non-coding DNA flanking each exon are sequenced for each of the genes listed below. Sequencing is accomplished by capturing specific regions with an optimized solution-based hybridization kit, followed by massively parallel sequencing of the captured DNA fragments. Additional Sanger sequencing is performed for any regions not captured or with insufficient number of sequence reads. All pathogenic, likely pathogenic, or variants of uncertain significance are confirmed by Sanger sequencing. Due to known ABCD1 pseudogenes, exons 7-10 of the ABCD1 gene are analyzed via Sanger sequencing.

Indications for Test

Individuals with clinical symptoms suggesting peroxisomal disorders.

Genes

Official Gene Symbol OMIM ID
ABCD1 300371
ABCD3 170995
ACOX1 609751
AGPS 603051
DNM1L 603850
GNPAT 602744
HSD17B4 601860
PEX1 602136
PEX10 602859
PEX11B 603867
PEX12 601758
PEX13 601789
PEX14 601791
PEX16 603360
PEX19 600279
PEX2 170993
PEX26 608666
PEX3 603164
PEX5 600414
PEX6 601498
PEX7 601757
PHYH 602026
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Diseases

Name Inheritance OMIM ID
Adrenoleukodystrophy 300100
Bile Acid Synthesis Defect, Congenital, 5 616278
D-Bifunctional Protein Deficiency 261515
Encephalopathy, Lethal, Due To Defective Mitochondrial And Peroxisomal Fission 614388
Peroxisomal Acyl-CoA Oxidase Deficiency 264470
Peroxisome biogenesis disorder 10A (Zellweger) 614882
Peroxisome biogenesis disorder 11A (Zellweger) 614883
Peroxisome biogenesis disorder 11B 614885
Peroxisome biogenesis disorder 12A (Zellweger) 614886
Peroxisome biogenesis disorder 13A (Zellweger) 614887
Peroxisome Biogenesis Disorder 14B 614920
Peroxisome biogenesis disorder 1A (Zellweger) 214100
Peroxisome biogenesis disorder 1B (NALD/IRD) 601539
Peroxisome biogenesis disorder 2A (Zellweger) 214110
Peroxisome biogenesis disorder 2B 202370
Peroxisome biogenesis disorder 3A (Zellweger) 614859
Peroxisome biogenesis disorder 3B 266510
Peroxisome biogenesis disorder 4A (Zellweger) 614862
Peroxisome biogenesis disorder 4B 614863
Peroxisome biogenesis disorder 5A (Zellweger) 614866
Peroxisome biogenesis disorder 5B 614867
Peroxisome biogenesis disorder 6A (Zellweger) 614870
Peroxisome biogenesis disorder 6B 614871
Peroxisome biogenesis disorder 7A (Zellweger) 614872
Peroxisome biogenesis disorder 7B 614873
Peroxisome biogenesis disorder 8A, (Zellweger) 614876
Peroxisome biogenesis disorder 8B 614877
Peroxisome Biogenesis Disorder 9B 614879
Refsum Disease, Classic 266500
Rhizomelic Chondrodysplasia Punctata Type 1 215100
Rhizomelic Chondrodysplasia Punctata Type 2 222765
Rhizomelic Chondrodysplasia Punctata, Type 3 600121

Related Tests

Name
Acyl-CoA Oxidase Deficiency via the ACOX1 Gene
Autism Spectrum Disorders and Intellectual Disability (ASD-ID) Comprehensive Panel
Comprehensive Fetal and Neonatal Loss Panel
Comprehensive Inherited Retinal Dystrophies (includes RPGR ORF15) Sequencing Panel
Congenital Bile Acid Synthesis Defect-5 via the ABCD3 Gene
Fetal Concerns Sequencing Panel
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the DNM1L Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX10 Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX11B Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX12 Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX13 Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX14 Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX16 Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX19 Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX1 Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX26 Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX2 Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX3 Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX5 Gene
Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum via the PEX6 Gene
Perrault Syndrome Type 1 via the HSD17B4 Gene
Refsum Disease via the PHYH Gene
Rhizomelic Chondrodysplasia Punctata Type 1 and Adult Refsum Disease via the PEX7 Gene
Rhizomelic Chondrodysplasia Punctata Type 2 via the GNPAT Gene
Rhizomelic Chondrodysplasia Punctata Type 3 via the AGPS Gene
Skeletal Disorders and Joint Problems Sequencing Panel
X-Linked Adrenoleukodystrophy via the ABCD1 Gene
X-Linked Intellectual Disability Sequencing Panel

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Braverman NE, Moser AB, Steinberg SJ. 2012. Rhizomelic Chondrodysplasia Punctata Type 1. In: Pagon RA, Adam MP, Ardinger HH, Bird TD, Dolan CR, Fong C-T, Smith RJ, and Stephens K, editors. GeneReviews(®), Seattle (WA): University of Washington, Seattle. PubMed ID: 20301447
  • Human Gene Mutation Database (Bio-base).
  • Itzkovitz B, Jiralerspong S, Nimmo G, Loscalzo M, Horovitz DDG, Snowden A, Moser A, Steinberg S, Braverman N. 2012. Functional characterization of novel mutations in GNPAT and AGPS, causing rhizomelic chondrodysplasia punctata (RCDP) types 2 and 3. Hum. Mutat. 33: 189–197. PubMed ID: 21990100
  • Jansen GA, Waterham HR, Wanders RJA. 2004. Molecular basis of Refsum disease: sequence variations in phytanoyl-CoA hydroxylase (PHYH) and the PTS2 receptor (PEX7). Hum. Mutat. 23: 209–218. PubMed ID: 14974078
  • Steinberg SJ, Moser AB, Raymond GV. 2012. X-Linked Adrenoleukodystrophy. In: Pagon RA, Adam MP, Ardinger HH, Bird TD, Dolan CR, Fong C-T, Smith RJ, and Stephens K, editors. GeneReviews(®), Seattle (WA): University of Washington, Seattle. PubMed ID: 20301491
  • Steinberg SJ, Raymond GV, Braverman NE, Moser AB. 2012. Peroxisome Biogenesis Disorders, Zellweger Syndrome Spectrum. In: Pagon RA, Adam MP, Bird TD, Dolan CR, Fong C-T, and Stephens K, editors. GeneReviewsTM, Seattle (WA): University of Washington, Seattle. PubMed ID: 20301621
  • Thai TP, Rodemer C, Jauch A, Hunziker A, Moser A, Gorgas K, Just WW. 2001. Impaired membrane traffic in defective ether lipid biosynthesis. Hum. Mol. Genet. 10: 127–136. PubMed ID: 11152660
  • Wanders RJA, Waterham HR. 2006. Peroxisomal disorders: the single peroxisomal enzyme deficiencies. Biochim. Biophys. Acta 1763: 1707–1720. PubMed ID: 17055078
  • Waterham HR, Ebberink MS. 2012. Genetics and molecular basis of human peroxisome biogenesis disorders. Biochim. Biophys. Acta 1822: 1430–1441. PubMed ID: 22871920
  • Waterham HR, Koster J, Roermund CWT van, Mooyer PAW, Wanders RJA, Leonard JV. 2007. A lethal defect of mitochondrial and peroxisomal fission. N. Engl. J. Med. 356: 1736–1741. PubMed ID: 17460227
  • Yoon G, Malam Z, Paton T, Marshall C, Hyatt E, Ivakine Z, Kemaladewi D, Forge C, Lee KS, Hawkins C, Cohn RD. 2014. G.O.6. Neuromuscular Disorders 24: 794. PubMed ID: Abstract
Order Kits
TEST METHODS

NextGen Sequencing using PG-Select Capture Probes

Test Procedure

We use a combination of Next Generation Sequencing (NGS) and Sanger sequencing technologies to cover the full coding regions of the listed genes plus ~20 bases of non-coding DNA flanking each exon.  As required, genomic DNA is extracted from the patient specimen.  For NGS, patient DNA corresponding to these regions is captured using an optimized set of DNA hybridization probes.  Captured DNA is sequenced using Illumina’s Reversible Dye Terminator (RDT) platform (Illumina, San Diego, CA, USA).  Regions with insufficient coverage by NGS are covered by Sanger sequencing.  All pathogenic, likely pathogenic, or variants of uncertain significance are confirmed by Sanger sequencing.

For Sanger sequencing, Polymerase Chain Reaction (PCR) is used to amplify targeted regions.  After purification of the PCR products, cycle sequencing is carried out using the ABI Big Dye Terminator v.3.0 kit.  PCR products are resolved by electrophoresis on an ABI 3730xl capillary sequencer.  In nearly all cases, cycle sequencing is performed separately in both the forward and reverse directions.

Patient DNA sequence is aligned to the genomic reference sequence for the indicated gene region(s). All differences from the reference sequences (sequence variants) are assigned to one of five interpretation categories, listed below, per ACMG Guidelines (Richards et al. 2015).

(1) Pathogenic Variants
(2) Likely Pathogenic Variants
(3) Variants of Uncertain Significance
(4) Likely Benign Variants
(5) Benign, Common Variants

Human Genome Variation Society (HGVS) recommendations are used to describe sequence variants (http://www.hgvs.org).  Rare variants and undocumented variants are nearly always classified as likely benign if there is no indication that they alter protein sequence or disrupt splicing.

Analytical Validity

As of March 2016, 6.36 Mb of sequence (83 genes, 1557 exons) generated in our lab was compared between Sanger and NextGen methodologies. We detected no differences between the two methods. The comparison involved 6400 total sequence variants (differences from the reference sequences). Of these, 6144 were nucleotide substitutions and 256 were insertions or deletions. About 65% of the variants were heterozygous and 35% homozygous. The insertions and deletions ranged in length from 1 to over 100 nucleotides.

In silico validation of insertions and deletions in 20 replicates of 5 genes was also performed. The validation included insertions and deletions of lengths between 1 and 100 nucleotides. Insertions tested in silico: 2200 between 1 and 5 nucleotides, 625 between 6 and 10 nucleotides, 29 between 11 and 20 nucleotides, 25 between 21 and 49 nucleotides, and 23 at or greater than 50 nucleotides, with the largest at 98 nucleotides. All insertions were detected. Deletions tested in silico: 1813 between 1 and 5 nucleotides, 97 between 6 and 10 nucleotides, 32 between 11 and 20 nucleotides, 20 between 21 and 49 nucleotides, and 39 at or greater than 50 nucleotides, with the largest at 96 nucleotides. All deletions less than 50 nucleotides in length were detected, 13 greater than 50 nucleotides in length were missed. Our standard NextGen sequence variant calling algorithms are generally not capable of detecting insertions (duplications) or heterozygous deletions greater than 100 nucleotides. Large homozygous deletions appear to be detectable.   

Analytical Limitations

Interpretation of the test results is limited by the information that is currently available.  Better interpretation should be possible in the future as more data and knowledge about human genetics and this specific disorder are accumulated.

When Sanger sequencing does not reveal any difference from the reference sequence, or when a sequence variant is homozygous, we cannot be certain that we were able to detect both patient alleles.  Occasionally, a patient may carry an allele which does not amplify, due to a large deletion or insertion.   In these cases, the report will contain no information about the second allele.  Our Sanger and NGS Sequencing tests are generally not capable of detecting Copy Number Variants (CNVs).

We sequence all coding exons for each given transcript, plus ~20 bp of flanking non-coding DNA for each exon.  Test reports contain no information about other portions of the gene, such as regulatory domains, deep intronic regions or any currently uncharacterized alternative exons.

In most cases, we are unable to determine the phase of sequence variants.  In particular, when we find two likely causative mutations for recessive disorders, we cannot be certain that the mutations are on different alleles.

Our ability to detect minor sequence variants due to somatic mosaicism is limited.  Sequence variants that are present in less than 50% of the patient’s nucleated cells may not be detected.

Runs of mononucleotide repeats (eg (A)n or (T)n) with n >8 in the reference sequence are generally not analyzed because of strand slippage during PCR.

Unless otherwise indicated, DNA sequence data is obtained from a specific cell-type (usually leukocytes from whole blood).   Test reports contain no information about the DNA sequence in other cell-types.

We cannot be certain that the reference sequences are correct.

Rare, low probability interpretations of sequencing results, such as for example the occurrence of de novo mutations in recessive disorders, are generally not included in the reports.

We have confidence in our ability to track a specimen once it has been received by PreventionGenetics.  However, we take no responsibility for any specimen labeling errors that occur before the sample arrives at PreventionGenetics.

Deletion/Duplication Testing Via Array Comparative Genomic Hybridization

Test Procedure

Equal amounts of genomic DNA from the patient and a gender matched reference sample are amplified and labeled with Cy3 and Cy5 dyes, respectively. To prevent any sample cross contamination, a unique sample tracking control is added into each patient sample. Each labeled patient product is then purified, quantified, and combined with the same amount of reference product. The combined sample is loaded onto the designed array and hybridized for at least 22-42 hours at 65°C. Arrays are then washed and scanned immediately with 2.5 µM resolution. Only data for the gene(s) of interest for each patient are extracted and analyzed.

Analytical Validity

PreventionGenetics' high density gene-centric custom designed aCGH enables the detection of relatively small deletions and duplications within a single exon of a given gene or deletions and duplications encompassing the entire gene. PreventionGenetics has established and verified this test's accuracy and precision.

Analytical Limitations

Our dense probe coverage may allow detection of deletions/duplications down to 100 bp; however due to limitations and probe spacing this cannot be guaranteed across all exons of all genes. Therefore, some copy number changes smaller than 100-300 bp within a targeted large exon may not be detected by our array.

This array may not detect deletions and duplications present at low levels of mosaicism or those present in genes that have pseudogene copies or repeats elsewhere in the genome.

aCGH will not detect balanced translocations, inversions, or point mutations that may be responsible for the clinical phenotype.

Breakpoints, if occurring outside the targeted gene, may be hard to define.

The sensitivity of this assay may be reduced when DNA is extracted by an outside laboratory.

Order Kits

Ordering Options


myPrevent - Online Ordering
  • The test can be added to your online orders in the Summary and Pricing section.
  • Once the test has been added log in to myPrevent to fill out an online requisition form.
REQUISITION FORM
  • A completed requisition form must accompany all specimens.
  • Billing information along with specimen and shipping instructions are within the requisition form.
  • All testing must be ordered by a qualified healthcare provider.

SPECIMEN TYPES
WHOLE BLOOD

(Delivery accepted Monday - Saturday)

  • Collect 3 ml -5 ml (5 ml preferred) of whole blood in EDTA (purple top tube) or ACD (yellow top tube). For Test #500-DNA Banking only, collect 10 ml -20 ml of whole blood.
  • For small babies, we require a minimum of 1 ml of blood.
  • Only one blood tube is required for multiple tests.
  • Ship blood tubes at room temperature in an insulated container. Do not freeze blood.
  • During hot weather, include a frozen ice pack in the shipping container. Place a paper towel or other thin material between the ice pack and the blood tube.
  • In cold weather, include an unfrozen ice pack in the shipping container as insulation.
  • At room temperature, blood specimen is stable for up to 48 hours.
  • If refrigerated, blood specimen is stable for up to one week.
  • Label the tube with the patient name, date of birth and/or ID number.

DNA

(Delivery accepted Monday - Saturday)

  • Send in screw cap tube at least 5 µg -10 µg of purified DNA at a concentration of at least 20 µg/ml for NGS and Sanger tests and at least 5 µg of purified DNA at a concentration of at least 100 µg/ml for gene-centric aCGH, MLPA, and CMA tests, minimum 2 µg for limited specimens.
  • For requests requiring more than one test, send an additional 5 µg DNA per test ordered when possible.
  • DNA may be shipped at room temperature.
  • Label the tube with the composition of the solute, DNA concentration as well as the patient’s name, date of birth, and/or ID number.
  • We only accept genomic DNA for testing. We do NOT accept products of whole genome amplification reactions or other amplification reactions.

CELL CULTURE

(Delivery preferred Monday - Thursday)

  • PreventionGenetics should be notified in advance of arrival of a cell culture.
  • Culture and send at least two T25 flasks of confluent cells.
  • Some panels may require additional flasks (dependent on size of genes, amount of Sanger sequencing required, etc.). Multiple test requests may also require additional flasks. Please contact us for details.
  • Send specimens in insulated, shatterproof container overnight.
  • Cell cultures may be shipped at room temperature or refrigerated.
  • Label the flasks with the patient name, date of birth, and/or ID number.
  • We strongly recommend maintaining a local back-up culture. We do not culture cells.
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