Screening For Lysosomal Storage Disorders Research Articles

Light and Shadows in Newborn Screening for Lysosomal Storage Disorders: Eight Years of Experience in Northeast Italy.

In the last two decades, the development of high-throughput diagnostic methods and the availability of effective treatments have increased the interest in newborn screening for lysosomal storage disorders. However, long-term follow-up experience is needed to clearly identify risks, benefits and challenges. We report our 8-year experience of screening and follow-up on about 250,000 neonates screened for four lysosomal storage diseases (Pompe disease, mucopolysaccharidosis type I, Fabry disease, Gaucher disease), using the enzyme activity assay by tandem mass spectrometry, and biomarker quantification as a second-tier test. Among the 126 positive newborns (0.051%), 51 infants were confirmed as affected (positive predictive value 40%), with an overall incidence of 1:4874. Of these, three patients with infantile-onset Pompe disease, two with neonatal-onset Gaucher disease and four with mucopolysaccharidosis type I were immediately treated. Furthermore, another four Gaucher disease patients needed treatment in the first years of life. Our study demonstrates the feasibility and effectiveness of newborn screening for lysosomal storage diseases. Early diagnosis and treatment allow the achievement of better patient outcomes. Challenges such as false-positive rates, the diagnosis of variants of uncertain significance or late-onset forms and the lack of treatment for neuronopathic forms, should be addressed.

An update on multiplexed mass spectrometry-based lysosomal storage disease diagnosis.

Lysosomal storage disorders (LSDs) are a type of inherited metabolic disorders in which biomolecules, accumulate as a specific substrate in lysosomes due to specific individual enzyme deficiencies. Despite the fact that LSDs are incurable, various approaches, including enzyme replacement therapy, hematopoietic stem cell transplantation, or gene therapy are now available. Therefore, a timely diagnosis is a critical initial step in patient treatment. The-state-of-the-art in LSD diagnostic uses, in the first stage, enzymatic activity determination by fluorimetry or by mass spectrometry (MS) with the aid of dry blood spots, based on different enzymatic substrate structures. Due to its sensitivity, high precision, and ability to screen for an unprecedented number of diseases in a single assay, multiplexed tandem MS-based enzyme activity assays for the screening of LSDs in newborns have recently received a lot of attention. Here, (i) we review the current approaches used for simultaneous enzymatic activity determination of LSDs in dried blood spots using multiplex-LC-MS/MS; (ii) we explore the need for designing novel enzymatic substrates that generate different enzymatic products with distinct molecular masses in multiplexed-MS studies; and (iii) we give examples of the relevance of affinity-MS technique as a basis for reversing undesirable immune-reactivity in enzyme replacement therapy.

Analysis of urinary oligosaccharide excretion patterns by UHPLC/HRAM mass spectrometry for screening of lysosomal storage disorders.

Oligosaccharidoses, sphingolipidoses and mucolipidoses are lysosomal storage disorders (LSDs) in which defective breakdown of glycan-side chains of glycosylated proteins and glycolipids leads to the accumulation of incompletely degraded oligosaccharides within lysosomes. In metabolic laboratories, these disorders are commonly diagnosed by thin-layer chromatography (TLC) but more recently also mass spectrometry-based approaches have been published. To expand the possibilities to screen for these diseases, we developed an ultra-high-performance liquid chromatography (UHPLC) with a high-resolution accurate mass (HRAM) mass spectrometry (MS) screening platform, together with an open-source iterative bioinformatics pipeline. This pipeline generates comprehensive biomarker profiles and allows for extensive quality control (QC) monitoring. Using this platform, we were able to identify α-mannosidosis, β-mannosidosis, α-N-acetylgalactosaminidase deficiency, sialidosis, galactosialidosis, fucosidosis, aspartylglucosaminuria, GM1 gangliosidosis, GM2 gangliosidosis (M. Sandhoff) and mucolipidosis II/III in patient samples. Aberrant urinary oligosaccharide excretions were also detected for other disorders, including NGLY1 congenital disorder of deglycosylation, sialic acid storage disease, MPS type IV B and GSD II (Pompe disease). For the latter disorder, we identified heptahexose (Hex7), as a potential urinary biomarker, in addition to glucose tetrasaccharide (Glc4), for the diagnosis and monitoring of young onset cases of Pompe disease. Occasionally, so-called "neonate" biomarker profiles were observed in young patients, which were probably due to nutrition. Our UHPLC/HRAM-MS screening platform can easily be adopted in biochemical laboratories and allows for simple and robust screening and straightforward interpretation of the screening results to detect disorders in which aberrant oligosaccharides accumulate.

The chitinases as biomarkers in immune-mediate diseases.

The role of chitinases has been focused as potential biomarkers in a wide number of inflammatory diseases, in monitoring active disease state, and predicting prognosis and response to therapies. The main chitinases, CHIT1 and YKL-40, are derived from 18 glycosyl hydrolases macrophage activation and play important roles in defense against chitin-containing pathogens and in food processing. Moreover, chitinases may have organ- as well as cell-specific effects in the context of infectious diseases and inflammatory disorders and able to induce tissue remodelling. The CHIT1 measurement is an easy, reproducible, reliable, and cost-effective affordable assay. The clinical use of CHIT1 for the screening of lysosomal storage disorders is quite practical, when proper cut-off values are determined for each laboratory. The potential of CHIT1 and chitinases has not been fully explored yet and future studies will produce many surprising discoveries in the immunology and allergology fields of research. However, since the presence of a null CHIT1 gene in a subpopulation would be responsible of false-negative values, the assay should be completed with the other markers such ACE and, if necessary, by genetic analysis when CHIT1 is unexpected low.

Establishment of Cutoff Values for Newborn Screening of Six Lysosomal Storage Disorders by Tandem Mass Spectrometry.

ObjectiveLysosomal storage disorders (LSDs) are becoming increasingly important in newborn screening, and tandem mass spectrometry (MS/MS) is widely used in newborn screening for LSDs through measurement of enzymatic activities in dried blood spots (DBSs). Overall, the determination of the cutoff value is important in such screening, and different laboratories have different methods of determining this value; most do not use a fixed cutoff value but rather calculate the corresponding batch cutoff value based on each batch of experimental data. In this study, we used MS/MS to screen for LSDs and sought to find an appropriate method to establish the cutoff value for LSD screening.MethodsA total of 38,945 samples from newborn blood tablets collected from various maternity hospitals in six cities in Shandong province, including Jinan, Dezhou, Heze, Linyi, Weifang, and Zibo, were tested using a Waters Xevo TQD tandem mass spectrometer; the experimental data were analyzed with MassLynx V4.1. The laboratory used 30% of the median GLA enzyme activity and 20% of the median ABG, ASM, GALC, IDUA, and GAA enzyme activities in every test as the cutoff values for that batch of experiments.ResultsThere were 254 suspicious positives in the initial screening test, including one case of Gaucher disease, one of Niemann-Pick disease, 47 of Krabbe disease, four of MPS-I, 21 of Fabry disease, and 180 of Pompe disease. After genetic screening, 11 children were diagnosed, including three with Pompe disease, three with Fabry disease, and five with Krabbe disease. In addition, the enzyme activity cutoff value of this experiment showed seasonal variation, which was initially believed to be related to the ambient temperature, such as the effect of ambient temperature on the human body or the temperature when the blood tablets dried naturally.ConclusionOverall, MS/MS can be used in LSD screening, and using different cutoff values in each batch of experiments is feasible. The ambient temperature might be a reason why the enzyme activity cutoff value has seasonal variation. More samples are needed to develop a method of determining cutoff values in laboratories.

The future of newborn screening for lysosomal disorders

The goal of newborn screening is to enhance the outcome of individuals with serious, treatable disorders through early, pre-symptomatic detection. The lysosomal storage disorders (LSDs) comprise a group of more than 50 diseases with a combined frequency of approximately 1:7000. With the availability of existing and new enzyme replacement therapies, small molecule treatments and gene therapies, there is increasing interest in screening newborns for LSDs with the goal of reducing disease-related morbidity and mortality through early detection. Novel screening methods are being developed, including efforts to enhance accuracy of screening using an array of multi-tiered, genomic, statistical, and bioinformatic approaches. While NBS data for Gaucher disease, Fabry disease, Krabbe disease, MPS I, and Pompe disease has demonstrated the feasibility of widespread screening, it has also highlighted some of the complexities of screening for LSDs. These include the identification of infants with later-onset, untreatable, and uncertain phenotypes, raising interesting ethical concerns that should be addressed as part of the NBS implementation process. Taken together, these efforts will provide critical, detailed data to help guide objective, ethically sensitive decision-making about NBS for LSDs.

Analysis of parent perception of newborn screening for lysosomal disorders

Lysosomal storage disorders (LSD) are a family of rare metabolic disorders that include Pompe disease and Mucopolysaccharidosis Type 1 (MPS1). Pompe disease is caused by deficiencies in the lysosomal enzyme alpha-glucosidase (GAA). This diagnosis is generally divided into infantile and late onset, but can exist along a spectrum. The main feature of Pompe disease is progressive muscle weakness, and in the infantile onset form, cardiomyopathy is present as well. MPS1 results from a deficiency in the lysosomal enzyme alpha-L-iduronidase (IDUA). Features include progressive neurological disease, skeletal abnormalities, cardiac disease, corneal clouding, hearing loss and hepatomegaly. The diagnosis has been stratified into MPS1 or attenuated MPS1, and severity or development of symptoms is dependent on the type. Both Pompe disease and MPS1 were recently added to the newborn screening panel. With their addition, a number of challenges have occurred including negative psychosocial impact on parents of infants who have an abnormal newborn screening result. The purpose of this study was to examine the experience and perceptions of parents/caregivers of an infant who had a positive newborn screening result for Pompe disease or MPS1. A survey was distributed via email and mailer, to parents and caregivers followed by the UPMC Children’s Hospital of Pittsburgh and the Children’s Hospital of Philadelphia. The survey consisted of questions specific to newborn screening of LSDs. Parent responses, were consistent with the literature, indicating that this period was categorized by stress and uncertainty. Of the five respondents, 100% described initial disclosure of the newborn screening results as difficult. Responses suggest that negative emotions were fueled by lack of information provided about the results, and provider’s lack of knowledge. Similar to published literature, this study indicated the majority of infants are not diagnosed with the most severe forms of the disorders. Results of this study may have implications for how genetic counselors care for these families and how they communicate with other providers. Likewise, it demonstrates public health relevance given the large scale in which newborn screening is utilized. This information can be used by providers in their own practice and may even influence standard of care.

Selective screening for lysosomal storage disorders in a large cohort of minorities of African descent shows high prevalence rates and novel variants.

Population studies point to regional and ethnicity‐specific differences in genetic predisposition for some lysosomal storage disorders (LSDs). The aim of the study was to determine the prevalence of the three treatable forms of lysosomal storage disorders (Gaucher disease [GD], Pompe disease [PD], and Fabry disease [FD]) in a cohort of mostly urban‐dwelling individuals of African ancestry, a previously unknown genetic landscape for LSDs. Large‐scale selective multistep biochemical and genetic screening was performed in patients seeking healthcare for various health concerns. Fluorimetric enzyme assays for GD, PD, and FD were performed on dried blood spots. Targeted gene sequencing was performed on samples that showed significantly lower enzyme activities (<10% of control mean) after two tiers of enzymatic screening. A total of 5287 unique samples representing a cross section of patients who visited Howard University Hospital and College of Medicine from 2015 to 2017 were included in the study. Study samples were obtained from a population where ~90% reported as African‐American, ~5% Hispanic, and <5% Caucasian or other. Regarding GD, three subjects had either homozygous or heterozygous mutations in the GBA gene. As to PD, eight subjects were either homozygous or compound heterozygous for GAA mutations, including three novel mutations: (a) c.472 A > G; p.T158A, (b) c.503G > T; p.R168L, (c) c.1985del. Regarding FD, two subjects had pathogenic GLA mutations, and four had single nucleotide polymorphisms in the 5'UTR, previously implicated in modulating gene expression. The findings highlight a higher incidence of abnormal enzyme levels and pathogenic mutations in the target population reflecting ancestry‐based specific genotype and phenotype variations.

A highly multiplexed biochemical assay for analytes in dried blood spots: application to newborn screening and diagnosis of lysosomal storage disorders and other inborn errors of metabolism

PurposeTo develop a multiplexed assay for the newborn screening of lysosomal storage disorders and additional inborn errors in a flexible, comprehensive, and affordable manner to keep up with the expansion of the newborn screening panel. MethodsUltraperformance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS) was chosen as the detection platform for its superiority compared to traditional flow-injection MS/MS. ResultsA high-throughput, 18-plex UPLC-MS/MS assay was developed for screening purposes with a sample turnaround time of 2.7minutes. The assay was consolidated such that only four dried blood spot punches were required, and it displayed good precision and reproducibility. ConclusionWe report a highly multiplexed UPLC-MS/MS assay that is appropriate for the newborn screening of 15 lysosomal storage diseases and 3 additional inborn errors. It can be further expanded to include additional conditions for which presymptomatic diagnosis may facilitate optimum treatment outcome.

Newborn Screening for Lysosomal Storage Disorders: Methodologies for Measurement of Enzymatic Activities in Dried Blood Spots.

All worldwide newborn screening (NBS) for lysosomal storage diseases (LSDs) is performed as a first-tier test by measurement of lysosomal enzymatic activities in dried blood spots (DBS). The currently two available methodologies used for measurement of enzymatic activities are tandem mass spectrometry (MS/MS) and digital microfluidics fluorimetry (DMF-F). In this chapter we summarize the workflows for the two platforms. Neither platform is fully automated, but the relative ease of workflow will be dependent upon the specific operation of each newborn screening laboratory on a case-by-case basis. We provide the screen positive rate (the number of below cutoff newborns per 100,000 newborns) from all NBS laboratories worldwide carrying out MS/MS-based NBS of one or more LSDs. The analytical precision of the MS/MS method is higher than that for DMF-F as shown by analysis of a common set of quality control DBS by the Centers for Disease Control and Prevention (CDC). Both the MS/MS and DMF-F platforms enable multiplexing of the LSD enzymes. An advantage of MS/MS over DMF-F is the ability to include assays of enzymatic activities and biomarkers for which no fluorimetric methods exist. Advantages of DMF-F over MS/MS are: (1) simple to use technology with same-day turn-around time for the lysosomal enzymes with the fastest rates compared to MS/MS requiring overnight analytical runs.; (2) the DMF-F instrumentation, because of its simplicity, requires less maintenance than the MS/MS platform.

Rapid and Modular Assembly of Click Substrates To Assay Enzyme Activity in the Newborn Screening of Lysosomal Storage Disorders.

Synthetic substrates play a pivotal role in the development of enzyme assays for medical diagnostics. However, the preparation of these chemical tools often requires multistep synthetic procedures complicating structural optimization and limiting versatility. In particular, substrates for enzyme assays based on tandem mass spectrometry need to be designed and optimized to fulfill the requirements to finally enable the development of robust diagnostic assays. In addition, isotope-labeled standards need to be prepared to facilitate accurate quantification of enzyme assay products. Here we report the development of a building block strategy for rapid and modular assembly of enzyme substrates using click chemistry as a key step. These click substrates are made up of a sugar moiety as enzyme responsive unit, a linker that can easily be isotope-labeled for the synthesis of internal standards, and a modifier compound that can readily be exchanged for structural optimization and analytical/diagnostic tuning. Moreover, the building block assembly eliminates the need for extensive optimization of different glycosylation reactions as it enables the divergent synthesis of substrates using a clickable enzyme responsive unit. The outlined strategy has been applied to obtain a series of synthetic α-l-iduronates and sulfated β-d-galactosides as substrates for assaying α-l-iduronidase and N-acetylgalactosamine-6-sulfate sulfatase, enzymes related to the lysosomal storage disorders mucopolysaccharidosis type I and type IVa, respectively. Selected click substrates were finally shown to be suitable to assay enzyme activities in dried blood spot samples from affected patients and random newborns.

Current State of the Art of Newborn Screening for Lysosomal Storage Disorders

Prospective full-population newborn screening for multiple lysosomal storage disorders (LSDs) is currently practiced in a few NBS programs, and several others are actively pursuing this course of action. Two platforms suitable for multiple LSD screening—tandem mass spectrometry (MS/MS) and digital microfluidic fluorometry (DMF)—are now commercially available with reagent kits. In this article, we review the methods currently used for prospective NBS for LSDs and objectively compare their workflows and the results from two programs in the United States that screen for the same four LSDs, one using MS/MS and the other DMF. The results show that the DMF platform workflow is simpler and generates results faster than MS/MS, enabling results reporting on the same day as specimen analysis. Furthermore, the performance metrics for both platforms while not identical, are broadly similar and do not indicate the superior performance of one method over the other. Results show a preponderance of inconclusive results for Pompe and Fabry diseases and for Hurler syndrome, due to genetic heterogeneity and other factors that can lead to low enzyme activities, regardless of the screening method. We conclude that either platform is a good choice but caution that post-analytical tools will need to be applied to improve the positive predictive value for these conditions.

Newborn Screening for Lysosomal Storage Disorders

Lysosomal storage disorders (LSDs) are a heterogeneous group of approximately 50 rare inherited metabolic conditions that result from enzyme deficiencies that interfere with lysosome function. Although often grouped together, there is great variability regarding age of onset, severity, treatment, and outcomes for each disorder and subtype. Currently, laboratory methods are available to test newborns for seven of these conditions. Although newborn screening programs remain state-based, each at a different phase of condition review and implementation, if newborn screening for LSDs has not yet been adopted by the state within which you practice, it likely will. Given the extremely low prevalence and limited provider familiarity with these conditions, this article provides an overview of LSDs and the seven conditions for which newborn screening is available. It offers information about each of the conditions including enzyme deficiency, mode of inheritance, incidence rates, types, clinical course, and available as well as potential treatment options.

Newborn Screening for Lysosomal Storage Disorders—Continuing Education Posttest

Newborn Screening for Lysosomal Storage Disorders—Continuing Education Posttest

Contribution of tandem mass spectrometry to the diagnosis of lysosomal storage disorders.

Tandem mass spectrometry (MS/MS) is a highly sensitive and specific technique. Thanks to the development of triple quadrupole analyzers, it is becoming more widely used in laboratories working in the field of inborn errors of metabolism. We review here the state of the art of this technique applied to the diagnosis of lysosomal storage disorders (LSDs) and how MS/MS has changed the diagnostic rationale in recent years. This fine technology brings more sensitive, specific, and reliable methods than the previous biochemical ones for the analysis of urinary glycosaminoglycans, oligosaccharides, and sialic acid. In sphingolipidoses, the quantification of urinary sphingolipids (globotriaosylceramide, sulfatides) is possible. The measurement of new plasmatic biomarkers such as oxysterols, bile acids, and lysosphingolipids allows the screening of many sphingolipidoses and related disorders (Niemann-Pick type C), replacing tedious biochemical techniques. Applied to amniotic fluid, a more reliable prenatal diagnosis or screening of LSDs is now available for fetuses presenting with antenatal manifestations. Applied to enzyme measurements, it allows high throughput assays for the screening of large populations, even newborn screening. The advent of this new method can modify the diagnostic rationale behind LSDs.

Newborn screening for lysosomal storage disorders by tandem mass spectrometry in North East Italy.

Lysosomal storage diseases (LSDs) are inborn errors of metabolism resulting from 50 different inherited disorders. The increasing availability of treatments and the importance of early intervention have stimulated newborn screening (NBS) to diagnose LSDs and permit early intervention to prevent irreversible impairment or severe disability. We present our experience screening newborns in North East Italy to identify neonates with Mucopolysaccharidosis type I (MPS I) and Pompe, Fabry, and Gaucher diseases. Activities of acid β-glucocerebrosidase (ABG; Gaucher), acid α-glucosidase (GAA; Pompe), acid α-galactosidase (GLA; Fabry), and acid α-L-iduronidase (IDUA; MPS-I) in dried blood spots (DBS) from all newborns during a 17-month period were determined by multiplexed tandem mass spectrometry (MS/MS) using the NeoLSD® assay system. Enzymatic activity cutoff values were determined from 3500 anonymous newborn DBS. In the screening study, samples were retested if the value was below cutoff and a second spot was requested, with referral for confirmatory testing and medical evaluation if a low value was obtained. From September 2015 to January 2017, 44,411 newborns were screened for the four LSDs. We recalled 40 neonates (0.09%) for collection of a second DBS. Low activity was confirmed in 20, who had confirmatory testing. Ten of 20 had pathogenic mutations: two Pompe, two Gaucher, five Fabry, and one MPS-I. The incidences of Pompe and Gaucher diseases were similar (1/22,205), with Fabry disease the most frequent (1/8882) and MPS-I the rarest (1/44411). The combined incidence of the four disorders was 1/4411 births. Simultaneously determining multiple enzyme activities by MS/MS, with a focus on specific biochemical markers, successfully detected newborns with LSDs. The high incidence of these disorders supports this screening program.

Heritability of hair cortisol and genetic overlap with psychological variables

Lysosomal storage disorders (LSDs) are a heterogeneous group of approximately 50 rare inherited metabolic conditions that result from enzyme deficiencies that interfere with lysosome function. Although often grouped together, there is great variability regarding age of onset, severity, treatment, and outcomes for each disorder and subtype. Currently, laboratory methods are available to test newborns for seven of these conditions. Although newborn screening programs remain state-based, each at a different phase of condition review and implementation, if newborn screening for LSDs has not yet been adopted by the state within which you practice, it likely will. Given the extremely low prevalence and limited provider familiarity with these conditions, this article provides an overview of LSDs and the seven conditions for which newborn screening is available. It offers information about each of the conditions including enzyme deficiency, mode of inheritance, incidence rates, types, clinical course, and available as well as potential treatment options.

Newborn Screening for Lysosomal Storage Disorders in Illinois: The Initial 15-Month Experience

To assess the outcomes of newborn screening for 5 lysosomal storage disorders (LSDs) in the first cohort of infants tested in the state of Illinois. Tandem mass spectrometry was used to assay for the 5 LSD-associated enzymes in dried blood spot specimens obtained from 219 973 newborn samples sent to the Newborn Screening Laboratory of the Illinois Department of Public Health in Chicago. The total number of cases with a positive diagnosis and the incidence for each disorder were as follows: Fabry disease, n = 26 (1 in 8454, including the p.A143T variant); Pompe disease, n = 10 (1 in 21 979); Gaucher disease, n = 5 (1 in 43 959); mucopolysaccharidosis (MPS) type 1, n = 1 (1 in 219 793); and Niemann-Pick disease type A/B, n = 2 (1 in 109 897). Twenty-two infants had a positive screen for 1 of the 5 disorders but could not be classified as either affected or unaffected after follow-up testing, including genotyping. Pseudodeficiencies for alpha-L-iduronidase and alpha-glucosidase were detected more often than true deficiencies. The incidences of Fabry disease and Pompe disease were significantly higher than published estimates, although most cases detected were predicted to be late onset. The incidences of Gaucher disease, MPS I, and Niemann-Pick disease were comparable with previously published estimates. A total of 16 infants could not be positively identified as either affected or unaffected. To validate the true risks and benefits of newborn screening for LSD, long term follow-up in these infants and those detected with later-onset disorders will be essential.

Effect of temperature on lysosomal enzyme activity during preparation and storage of dried blood spots.

The use of dried blood spots (DBS) for the assay of lysosomal enzymes has facilitated the implementation of pilot studies for newborn screening for lysosomal storage disorders in various developed countries. The aim of the study was to determine the influence of ambient temperature during DBS preparation and storage on lysosomal enzyme activity in a developing, tropical country. Blood samples from 12 healthy subjects collected on a S&S 903 filter paper were dried and stored at different temperatures for different periods of time. Activities of five lysosomal enzymes (acid α-glucosidase, acid α-galactosidase, acid β-glucocerebrosidase, acid sphingomyelinase, and galactocerebrosidase) were determined by tandem mass spectrometric and fluorimetric (acid α-glucosidase and acid β-glucocerebrosidase only) assays. The mean activities of all five enzymes decreased significantly when DBS was dried at temperatures above 24°C (P<.0001). DBS stored at 4°C, 24°C, 30°C, 37°C, and 45°C for 10days and more, also showed significant reduction in activities of all five enzymes (P<.0001). The results highlight the importance of maintaining the correct ambient temperature during DBS preparation and storage to avoid false positive results when screening for lysosomal storage disorders.

Newborn Screening for Krabbe Disease and Other Lysosomal Storage Disorders: Broad Lessons Learned

Newborn screening (NBS) for Krabbe disease (KD) began in New York (NY) in August 2006. In summary, after eight years of screening there were five infants identified with early-onset Krabbe disease. Four underwent transplant, two are surviving with moderate to severe handicaps, and two died from transplant-related complications. An additional forty-six asymptomatic infants were found to be at moderate or high risk for disease. Screening for KD is both analytically and medically challenging; since screening for KD possesses both of these challenges, and many more, the lessons learned thus far could be used to predict the challenges that may be faced when screening for other lysosomal storage disorders (LSDs). This paper briefly reviews reports of NBS for LSDs from varied world programs. The challenges encountered in screening for KD in NY will be highlighted, and this experience, combined with hindsight, will inform what may be expected in the future as screening for LSDs expands.